David L. Knight

Paroxetine binding to the rat norepinephrine transporter in vivo

BACKGROUND The norepinephrine transporter (NET)/uptake site is an antidepressant-sensitive transporter located on plasma membranes of noradrenergic neurons and other specialized cells that remove norepinephrine (NE) from the synapse to terminate the actions of NE. The antidepressant paroxetine is believed to produce its therapeutic effects primarily by acting as a highly selective antagonist of the serotonin transporter (SERT). However, in vitro data indicates that paroxetine inhibits the NET. The present study was designed to determine whether paroxetine inhibits in NET in vivo. METHODS Rats were administered paroxetine (6.5, 10.0, or 15.0 mg/kg/day) via osmotic minipumps for 1 week. Following attainment of steady state serum concentrations, cortical NET function was assessed by both [3H]-nisoxetine binding and [3H]-norepinephrine uptake assays conducted ex vivo. RESULTS In unwashed brain homogenates, serum paroxetine concentrations greater than 100 ng/mL were positively correlated with the observed Kd for [3H]-nisoxetine. At [3H]-nisoxetine concentrations associated with 50% transporter occupancy in vehicle treated rats, [3H]-nisoxetine binding was decreased 21% and 34% in rats exhibiting serum paroxetine concentrations > 100 ng/mL and > 500 ng/mL, respectively. CONCLUSIONS Although paroxetine is a very potent inhibitor of the SERT, paroxetine also inhibits the NET at serum concentrations > 100 ng/mL. This novel finding may underlie the broad therapeutic utility of paroxetine in mood and anxiety disorders.

Effects of Sodium Valproate on Corticotropin-Releasing Factor Systems in Rat Brain

We hypothesized that divalproex sodium, an anticonvulsant effective in the acute treatment of mania, may act upon neuropeptide systems that utilize corticotropin-releasing factor (CRF). Pharmacokinetic studies demonstrated that valproate has an apparent elimination half life of 17 minutes in rats after acute administration and that there is a nonlinear relationship between chronic dose and serum drug concentration. Acute valproate treatment neither altered plasma adrenocorticotropic hormone (ACTH) or corticosterone concentrations nor produced changes in CRF concentration in any of 10 brain regions examined. Subchronic treatment via SC-implanted osmotic minipumps (875 mg/kg/day × 7 days) resulted in decreased CRF concentrations in the median eminence and raphe nuclei. Moreover, CRF mRNA expression was decreased in the central nucleus of the amygdala (CeA) and paraventricular nucleus (PVN) of the hypothalamus. The benzodiazepine alprazolam, also a positive modulator of GABAergic function, similarly decreases CRF mRNA expression in the CeA. These results suggest that the mood stabilizing effects of valproic acid may be mediated in part by alterations in CRF neuronal activity.

The simultaneous determination of neurotensin and its major fragments by on-line trace enrichment HPLC with electrochemical detection.

An HPLC assay using on-line cation exchange trace enrichment and acetonitrile gradient elution, ion pair reverse phase separation with electrochemical detection (EC) is described for the simultaneous determination of the tridecapeptide neurotensin (NT) and six of its fragments. Cyclic voltammetric analysis indicated that the oxidative electrochemical properties of NT and its fragments is not merely a function of the sum of its electroactive amino acids (i.e. tyrosine) but reflects the presence and association of other amino acids (e.g. the arginine-arginine pair at position 8-9). Using the described method, NT1-6, NT1-8, NT1-10, NT1-11, NT8-13, NT9-13 and NT1-13 were baseline resolved within 20 min with a limit of detection varying from 1 to 5 ng peptide/injection. Other structurally similar or quantitatively significant neuropeptides (e.g. substance P, somatostatin, bombesin) did not interfere. Initial application of this on-line trace enrichment HPLC-EC assay to the question of the molecular nature of NT in unprocessed human CSF indicated the predominance of NT1-13 with an apparent formation of NT1-8 and NT9-13 resulting from more vigorous sample preparation techniques. The improvements in assay specificity, signal-to-noise ratios, biomatrix compatibility and assayable sample volume compared to non-enrichment HPLC-EC are discussed.