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Serotonin transport is modulated differently by tetanus toxin and growth factors
Carles Gil1, Abderrahim Najib, José Aguilera
1Departament de Bioquímica i de Biologia Molecular, Institut de Neurociències, Universitat Autònoma de Barcelona, E-08193 Cerdanyola del Vallés, Barcelona, Spain.
Abstract:
It has been previously shown that 5-HT uptake inhibition produced by tetanus toxin (TeTx) corresponds to a non-competitive inhibition, and it is preceded by phosphorylation of the tyrosine-kinase receptor trkA, phospholipase C activation and translocation of protein kinase C isoforms [FEBS Lett. 481 (2000) 177; FEBS Lett. 486 (2000) 136]. In the present work, it is shown that agonists of tyrosine-kinase receptors (NGF, EGF, basic FGF) enhance Na(+)-dependent, 5-hydroxytryptamine (serotonin, 5-HT) uptake in the synaptosomal-enriched P(2) fraction from rat-brain, suggesting a divergence in the intracellular signal pathways triggered by TeTx and by agonists of TyrK receptors. Co-applications of TeTx and agonists of TyrK receptors result in a mutual and partial reversion of their effects on 5-HT transport. In spite of their differences on transport, TeTx, TPA and NGF produce an increase in serotonin transporter phosphorylation in Ser separately, which is abolished by the PKC-inhibitor bisindolylmaleimide-1. Co-application of sodium vanadate, a tyrosine-phosphatase inhibitor, partially abolishes the effect produced by TeTx, whereas genistein, a tyrosine-kinase inhibitor, does not exert any variation of TeTx inhibition. Analyses by immunoblotting of the activation of specific PKC isoforms activation, determined as translocation to the membrane compartment, reveals differences in the pattern produced by NGF and TeTx. PKC gamma, delta, and epsilon isoforms are equally activated by both compounds, whereas the beta isoform is activated in a sustained manner only by TeTx, and the alpha isoform is only down-regulated by NGF. The aim of the present work was to explore whether NGF have the same effect on 5-HT transport than TeTx, since both compounds share the ability of activate part of the same transduction pathways. In spite of this, growth factors and TeTx show an opposite effect on 5-HT transport, even though SERT phosphorylation is enhanced in both cases. The differential effect on alpha- and beta-PKC isoenzymes found between NGF and TeTx action could explain this apparent discrepancy.
Insights
Tetanus toxin (TeTx) inhibits serotonin uptake, while growth factors like NGF enhance it, despite both increasing serotonin transporter phosphorylation. Differential activation of protein kinase C (PKC) isoforms by TeTx and NGF explains this opposite effect on serotonin transport.
Area of Science:
- Neuroscience
- Molecular Biology
- Biochemistry
Background:
- Tetanus toxin (TeTx) inhibits serotonin (5-HT) uptake via non-competitive inhibition, involving tyrosine-kinase receptor (TrkA) phosphorylation, phospholipase C activation, and protein kinase C (PKC) translocation.
- Tyrosine-kinase receptor agonists, such as nerve growth factor (NGF), epidermal growth factor (EGF), and basic fibroblast growth factor (bFGF), are known to influence cellular signaling pathways.
Purpose of the Study:
- To investigate whether NGF elicits the same effect on serotonin transport as TeTx, given their shared ability to activate similar transduction pathways.
- To elucidate the divergent intracellular signaling mechanisms underlying the effects of TeTx and TyrK receptor agonists on serotonin uptake.
Main Methods:
- Utilized synaptosome-enriched P(2) fraction from rat brain to study Na(+)-dependent 5-HT uptake.
- Employed co-application experiments with TeTx and TyrK receptor agonists.
- Analyzed serotonin transporter (SERT) phosphorylation using immunoblotting and PKC isoform activation via membrane translocation analysis.
- Investigated the roles of tyrosine-phosphatase and tyrosine-kinase inhibitors (sodium vanadate and genistein, respectively).
Main Results:
- NGF, EGF, and bFGF enhanced Na(+)-dependent 5-HT uptake, contrasting with TeTx-induced inhibition.
- Co-application of TeTx and TyrK agonists resulted in partial mutual reversion of their effects on 5-HT transport.
- Both TeTx and NGF increased SERT phosphorylation, an effect abolished by a PKC inhibitor.
- Differential activation patterns of PKC isoforms were observed: TeTx sustainedly activated PKC beta, while NGF down-regulated PKC alpha. PKC gamma, delta, and epsilon were activated by both.
Conclusions:
- Despite shared signaling pathway activation and increased SERT phosphorylation, TeTx and growth factors exert opposing effects on 5-HT transport.
- The differential modulation of PKC alpha and beta isoforms by TeTx and NGF provides a molecular basis for their contrasting effects on serotonin transport.