Fenfluramine‐induced serotonergic neurotoxicity in mice: lack of neuroprotection by inhibition/ablation of nNOS

Abstract

Previous studies have implicated a role for nitric oxide (NO) and peroxynitrite in methamphetamine-induced dopaminergic neurotoxicity. The present study was undertaken to investigate whether NO is involved in serotonergic neurotoxicity caused by fenfluramine. In the first experiment, the effect of the neuronal nitric oxide synthase (nNOS) inhibitor 7-nitroindazole (7-NI; 25 mg/ kg · 4) on fenfluramine (25 mg/kg · 4)-induced serotonergic neurotoxicity in Swiss Webster mice was investigated. In the second experiment, the effect of fenfluramine (25 mg/kg · 4) on nNOS (-/-) and wild-type (WT) mice was investigated. Fenfluramine induced hypothermia in all three mouse strains, and 7-NI had no thermoregulatory effect. Selective depletion of 5-HT and 5-HT transporter binding sites in the striatum, frontal cortex and hippocampus in all three mouse strains was observed, with no evidence of dopaminergic neurotoxicity. In the first experiment, 7-NI did not attenuate serotonergic neurotoxicity in Swiss Webster mice. In the second experiment, nNOS(-/-) and WT mice were equally sensitive to serotonergic neurotoxicity. These findings suggest that NO and peroxynitrite do not mediate fenfluramineinduced serotonergic neurotoxicity, and that NO is a selective mediator of amphetamines-induced dopaminergic neurotoxicity. Keywords: 7-nitroindazole, fenfluramine, methamphetamine, nitric oxide, serotonergic and dopaminergic neurotoxicity. Fenfluramine is an amphetamine analog with anorectic properties that inhibits the reuptake and increases the release of 5-hydroxytryptamine (5-HT; serotonin). High doses of fenfluramine cause long-lasting depletion of 5-HT and 5-HT transporter (5-HTT) binding sites (Kleven and Seiden 1989;O'Callaghan and Miller 1994), deficits that could be consistent with neurotoxic insult in rodents and non-human primates (Schuster et al. 1986;Ricaurte et al. 1991;McCann et al. 1994). The mechanism underlying the development of serotonergic neurotoxicity is unclear. Endogenous formation of 5,6-dihydroxytryptamine due to 5-HT excess release following the administration of substituted amphetamines (Commins et al. 1987), and the generation of other products of 5-HT oxidation (Wrona et al. 1995) may be involved in degeneration of 5-HT nerve terminals.Other amphetamine analogs, such as methamphetamine (METH) and 3,4-methylenedioxy-methamphetamine (MDMA) cause varying degrees of dopaminergic and serotonergic neurotoxicity, depending on the species examined. Evidence suggests the involvement of free radicals and, specifically, reactive oxygen species in amphetamines-induced dopaminergic neurotoxicity (Cadet and Brannock 1998;Hanson et al. 1998). Peroxynitrite, formed from the interaction between nitric oxide (NO) and superoxide radicals, is considered a major neurotoxin (Beckman et al. 1990). Inactivation of tyrosine hydroxylase (Kuhn et al. 1999), the dopamine transporter (DAT; Park et al. 2002), and oxidation of dopamine (LaVoie and Hastings 1999) by peroxynitrite are thought to be involved in METH-induced dopamin...

Differential Effects of Amphetamines-Induced Neurotoxicity on Appetitive and Aversive Pavlovian Conditioning in Mice

“…The amphetamine doses chosen were based on our previous studies on the neurotoxic effects of METH (Itzhak and Ali, 1996;Itzhak et al, 2002); FEN (Itzhak et al, 2003b), and MDMA (Itzhak et al, 2003a) in Swiss Webster mice. Control mice received saline (0.9% NaCl) injections.…”

“…We have previously shown that METH and FEN selectively deplete dopaminergic (Itzhak and Ali, 1996) and serotonergic (Itzhak et al, 2003b) markers, respectively, while MDMA reduces both DA and 5-HT markers in Swiss Webster mice (Itzhak et al, 2003a;Itzhak and AchatMendes, 2004). Using these mouse models, we investigated the effect of amphetamines-induced injury to individual monoamine systems on CPP and CPA.…”

Methamphetamine and MDMA (Ecstasy) Neurotoxicity: 'of Mice and Men'

“…For instance, administration of METH (5 mg kg 71 63; 3 h apart) to Swiss Webster mice caused selective depletion of striatal DA, DA metabolites 3,4-dihydroxyphenylacetic acid (DO-PAC), homovanillic acid (HVA) and DAT binding sites, with no evidence of 5-HT depletion (26). Administration of fenfluramine (25 mg kg 71 62; 6 h apart; for 2 days) caused selective depletion of 5-HT and 5-HT transporter (5-HTT) binding sites in striatum, frontal cortex and hippocampus with no evidence of dopaminergic neurotoxicity (27). However, administration of MDMA (15 mg kg 71 62; 8 h apart, for 2 days) caused marked depletion of striatal DA and DAT binding sites as well as depletion of 5-HT in the striatum and hippocampus, and depletion of 5-HTT binding sites in the frontal cortex.…”

“…To test this hypothesis we investigated whether blockade of nNOS or ablation of nNOS (KO mice) provides protection against serotonergic neurotoxicity induced by the amphetamine analog fenfluramine (27). We found that neither inhibition of nNOS by 7-NI in Swiss Webster mice nor the absence of the nNOS gene in the nNOS (7/7) mice afforded protection against serotonergic neurotoxicity caused by high doses of fenfluramine (25 mg kg 71 62; for 2 days).…”

Serotonin neurotoxins — past and present

“…MDMA (single or repeated doses) had no effect on striatal 5-HT or 5-HIAA levels. The same group of authors (Itzhak et al, 2003) has reported that nNOS inactivation (by gene knockout) or submaximal inhibition of nNOS activity by a high dose of 7-nitroindazole (25 mg/kg i.p.) failed to protect central serotonergic systems in Swiss Webster mice against the long-term 5-HT depleting effect of a high-dose treatment regimen with d,l-Fen (25 mg/kg i.p., twice daily for two days) suggesting that NO-dependent mechanisms have no major role in the serotonergic actions of Fen.…”