Endogenous Release of Neuronal Serotonin and 5‐Hydroxyindoleacetic Acid in the Caudate‐Putamen of the Rat as Revealed by Intracerebral Dialysis Coupled to High‐Performance Liquid Chromatography with Fluorimetric Detection

Abstract: Extracellular levels of endogenous serotonin (5‐HT) and its major metabolite, 5‐hydroxyindoleacetic acid (5‐HIAA), were measured in the caudate‐putamen of anesthetized and awake rats using intracerebral microdialysis coupled to HPLC with fluorimetric detection. A dialysis probe (of the loop type) was perfused with Ringer solution at 2 μl/min, and samples collected every 30 or 60 min. Basal indole levels were followed for up to 4 days in both intact and 5,7‐dihydroxytryptamine (5,7‐DHT) lesioned animals. Immediately after the probe implantation, the striatal 5‐HT levels were about 10 times higher than the steady‐state levels that were reached after 7‐8 h of perfusion. The steady‐state baseline levels, which amounted to 22.5 fmol/30 min sampling time, remained stable for 4 days. In 5,7‐DHT‐denervated animals, the steady‐state levels of 5‐HT, measured during the second day after probe implantation, were below the limit of detection (<10 fmol/60 min). However, during the first 6h post‐implantation, the 5‐HT output was as high as in intact animals, which suggests that the high 5‐HT levels recovered in association with probe implantation were blood‐derived. As a consequence, all other experiments were started after a delay of at least 12 h after implantation of the dialysis probe. In awake, freely moving animals, the steady‐state 5‐HT levels were about 60% higher than in halothane‐anesthetized animals, whereas 5‐HIAA was unaffected by anesthesia. KCI (60 and 100 mM) added to the perfusion fluid produced a sharp increase in 5‐HT output that was eight‐fold at the 60 mM concentration and 21‐fold at the 100 mM concentration. In contrast, 5‐HIAA output dropped by 43 and 54%, respectively. In 5,7‐DHT‐lesioned animals, the KCl‐evoked (100 mM) release represented less than 5% of the peak values obtained for the intact striata. Omission of Ca2+ from the perfusion fluid resulted in a 70% reduction in baseline 5‐HT output, whereas the 5‐HIAA levels remained unchanged. High concentrations of tetrodotoxin (TTX) added to the perfusion medium (5‐50 μM) resulted in quite variable results. At a lower concentration (1 μM), however, TTX produceda 50% reduction in baseline 5‐HT release, whereas the 5‐HIAA output remained unchanged. The 5‐HT reuptake blocker, indalpine, increased the extracellular levels of 5‐HT sixfold when added to the perfusion medium (1 μM), and threefold when given intraperitoneally (5 mg/kg). By contrast, the 5‐HIAA level remained unaffected during indalpine infusion. Application of TTX (1 μM) under simultaneous 5‐HT uptake blockade induced a decrease in 5‐HT output by 62–71%. p‐Chloroamphetamine (2.5 mg/kg, i.p.) induced a 12‐fold increase in 5‐HT release and reduced the 5‐HIAA output by about 50%. The p‐chloroamphetamine‐induced increase in 5‐HT release was 10 times lower in the 5,7‐DHT‐denervated striatum. Pargyline (75 mg/kg, i.p.) increased the extracellular levels of 5‐HT 11‐fold within 6 h, and reduced the 5‐HIAA levels by 80%. The 5‐HT receptor agonist, 5‐methoxy‐N,N‐dimethyltryptamine (1 mg/kg, i.p.), produced an immediate reduction of about 50% in 5‐HT release and a small (11 %) decrease in 5‐HIAA output. It is concluded (1) that intracerebral microdialysis coupled to HPLC with fluorimetric detection provides a useful method for the study of extracellular 5‐HT and 5‐HIAA levels; (2) that steady‐state levels of 5‐HT and 5‐HIAA recovered in the dialysis perfusate are neuronally derived, but these steady‐state levels are reached only after a minimum of 7–8 h after probe implantation; (3) that changes in striatal extracellular levels of 5‐HT are closely related to changes in serotonergic synaptic activity; and (4) that extracellular levels of 5‐HIAA are a poor indicator of synaptic activity, and instead primarily reflect intraneuronal metabolism.