The major finding of the present work is that endogenous 5-HT, released by serotonergic terminals upon the action of MDMA in the CA1 region of hippocampal slices, increases the magnitude of LTP induced by TB5 stimulation at the CA3/CA1 synapses.

MDMA effects on LTP are mediated by release of endogenous serotonin

We investigated the effects of endogenous 5-HT released by MDMA on LTP induced by a pattern of afferent stimulation that in vitro appears to be optimal for producing LTP (Larson and Lynch 1986; Otto et al. 1991; Pugliese et al. 1994). We monitored the increase in synaptic transmission induced by TB5 stimulation with recordings from the stratum radiatum, where the fEPSP closely reports the changes in synaptic strength at CA3/CA1 dendritic synapses (Johnston and Wu 1995). We found that endogenous 5-HT released by MDMA facilitated LTP of fEPSP recorded at the level of CA3/CA1 synapses.

In a recent work, Rozas et al. (2012) found an increase in LTP following the application of a high concentration (50 µM) of MDMA. At this concentration, MDMA releases 5-HT, noradrenaline and dopamine (Fitzgerald and Reid 1993). Although the Authors ascribed part of MDMA action to dopamine release, the specific contribution of the three monoamines to the effect of MDMA on LTP was not characterized and the net effect of endogenous 5-HT on LTP remained unknown.

The present work was specifically directed to investigate the effect of endogenous 5-HT by using a low concentration (10 µM) of MDMA in the presence of nisoxetine, a protocol shown to selectively produce serotonergic effects in the CA1 hippocampus with its maximal effects at the time point (15 min) that we chose for LTP induction (Mlinar et al. 2008). The facilitation of LTP produced by MDMA persisted in the presence of nisoxetine and was prevented by citalopram that selectively blocks the entry of MDMA in 5-HT terminals and reversal of 5-HT transporter (Rudnick and Wall 1992; Sitte et al. 2000), thereby preventing 5-HT release. Collectively, our data show that the facilitatory effect of MDMA on fEPSP LTP was mediated by the release of endogenous 5-HT and that endogenous noradrenaline and dopamine did not contribute to this effect.

When considered in the context of MDMA effects, our results indicate that a substantial amount of the facilitation of LTP exerted by the drug is mediated by the 5-HT released and that an impairment of LTP-dependent hippocampal processing needs not to be assumed to explain drug-induced mild cognitive impairment observed in humans (Kuypers and Ramaekers 2007; Stough et al. 2012) and laboratory animals (Moyano et al. 2004; Rodsiri et al. 2011; but see Abad et al. 2014). The action on other, non-hippocampal and/or non-serotonergic, mechanisms of cognitive processing, e.g., in the prefrontal cortex (see Jensen and Lisman 2005), likely underlies the mild cognitive impairment produced by acute administration of MDMA.

Effects of endogenous versus exogenous 5-HT on hippocampal neurotransmission and plasticity

The effects of 5-HT on LTP have been relatively poorly studied, and there is little convergence among investigations carried out in vitro and in vivo, on the outcome of 5-HT modulatory effects on LTP. In the dentate gyrus in vivo (Bliss et al. 1983), selective depletion of serotonin reduces the magnitude of LTP and electrical activation of median raphe nucleus; hence, release of 5-HT, increases LTP (Klancnik and Phillips 1991). Furthermore, lesions of the serotonergic system impair LTP which is restored by raphe grafts together with part of the behavioral correlates of hippocampal activity (Richter-Levin and Segal 1991). These in vivo results suggest that 5-HT exerts a facilitatory modulation of LTP and hippocampal encoding activity. In contrast, the results obtained in slices with the application of exogenous 5-HT (Corradetti et al. 1992, Villani and Johnston 1993; Staubli and Otaky 1994) have led to the widely accepted view that 5-HT blocks hippocampal LTP (Bliss et al. 2007).

The present results show that endogenous 5-HT exerts a facilitating effect on LTP at the CA3/CA1 synapses which is the opposite of that described for exogenous 5-HT (Staubli and Otaky 1994).

Although one possible explanation for the block of synaptic LTP found by Staubli and Otaky (1994) may reside in the use of a high concentration (260 µM) of 5-HT, these results seem inscribed in a wider discrepancy between the findings obtained with exogenous and endogenous 5-HT on hippocampal neurotransmission.

Similar to what happens with LTP, contrasting effects of exogenous and endogenous 5-HT are found for normal neurotransmission in the CA1 hippocampus in vitro. Notably, the predominant effect of exogenous 5-HT application is the 5-HT 1A receptor-mediated hyperpolarization of pyramidal neurons and subsequent inhibition of neurotransmission (Jahnsen 1980; Segal 1980; Ropert 1988; Pugliese et al. 1998), whereas endogenous 5-HT released in vitro from 5-HT terminals produces an initial inhibitory effect followed by a marked increase in pyramidal neuron excitability resulting in enhancement of neurotransmission (Mlinar et al. 2008).

In this context, it should be considered that in slices, 5-HT 1B receptors expressed by CA1 pyramidal neurons appear fully activated by nanomolar concentrations of 5-HT as well as by endogenous 5-HT released by MDMA (Mlinar and Corradetti 2003) or fenfluramine (Winterer et al. 2011), showing that endogenous 5-HT is released at concentrations appropriate for activating 5-HT 1B receptors. On the other hand, micromolar concentrations of exogenous 5-HT are currently used to reveal its hyperpolarizing effect through 5-HT 1A receptors although 5-HT has similar, low nanomolar, affinity for 5-HT 1A and 5-HT 1B receptors. It is conceivable that the established use of high concentrations of exogenous 5-HT in slices, required to detect in somatic intracellular recordings the hyperpolarizing effect of 5-HT 1A receptors located primarily in distal dendrites, results in a disproportional activation of 5-HT 1A receptors compared with endogenous 5-HT. Furthermore, endogenous 5-HT is released by serotonergic terminals and therefore respects their distribution within the CA1 region, which likely results in a different pattern of 5-HT receptor activation compared to exogenous 5-HT.

Functional implications of LTP facilitation

Our data do not resolve the question of whether 5-HT is required or not for physiological LTP in vivo, but clearly show that a sustained increase in 5-HT at a synaptic level does not impair the cellular mechanism(s) required for strengthening synaptic transmission between CA3 and CA1 pyramidal neurons.

From this perspective, our data appear to reconcile the in vitro effects of 5-HT on LTP with those apparently exerted by SSRIs that enhance 5-HT neurotransmission, increase hippocampal activity in vivo (McKie et al. 2005; Anderson et al. 2011) and have positive memory and behavioral effects in healthy humans (Harmer et al. 2002, 2003), whereas the converse occurs with acute tryptophan depletion (Hayward et al. 2005; van der Veen et al. 2006). In fact, hippocampal synaptic LTP is fundamental for learning and memory and its disruption by SSRI-induced enhancement of 5-HT signaling would be expected to result in a remarkable impairment of these functions in vivo.

5-hydroxytryptamine neurotransmission is also implicated in emotional (Del-Ben et al. 2008) and social processing (Young and Leyton 2002), as well as in affiliative behavior (Tse and Bond 2002). The here-reported 5-HT-induced enhancement of LTP at CA3/CA1 synapses of pyramidal cells likely translates into facilitated learning-mediated hippocampal activity and emotional processing in vivo. Consistent with this view, positive social stimuli activate the hippocampus in normal subjects, but not in depressed patients (Schaefer et al. 2006), and sustained hippocampal activation occurs during a period of subjective emotion (Garrett and Maddock 2006).

In conclusion, our finding that endogenous 5-HT does not inhibit, but facilitates LTP in vitro suggests a strengthening of hippocampal encoding linked to learning, memory and emotional processes which appears more consistent with the effects observed in vivo following 5-HT system manipulations. Since the release of 5-HT produced by MDMA appears to preserve and facilitate hippocampal encoding through LTP, it is conceivable that 5-HT contributes to SSRI antidepressant effects in humans such as positive mood changes, cognitive improvement, enhanced empathy and increased social interaction.