Effects of escitalopram on synaptic density in the healthy human brain: a randomized controlled trial

Selective serotonin reuptake inhibitors (SSRIs) are widely used for treating neuropsychiatric disorders. However, the exact mechanism of action and why effects can take several weeks to manifest is not clear. The hypothesis of neuroplasticity is supported by preclinical studies, but the evidence in humans is limited. Here, we investigate the effects of the SSRI escitalopram on presynaptic density as a proxy for synaptic plasticity. In a double-blind placebo-controlled study (NCT04239339), 32 healthy participants with no history of psychiatric or cognitive disorders were randomized to receive daily oral dosing of either 20 mg escitalopram (n = 17) or a placebo (n = 15). After an intervention period of 3–5 weeks, participants underwent a [11C]UCB-J PET scan (29 with full arterial input function) to quantify synaptic vesicle glycoprotein 2A (SV2A) density in the hippocampus and the neocortex. Whereas we find no statistically significant group difference in SV2A binding after an average of 29 (range: 24–38) days of intervention, our secondary analyses show a time-dependent effect of escitalopram on cerebral SV2A binding with positive associations between [11C]UCB-J binding and duration of escitalopram intervention. Our findings suggest that brain synaptic plasticity evolves over 3–5 weeks in healthy humans following daily intake of escitalopram. This is the first in vivo evidence to support the hypothesis of neuroplasticity as a mechanism of action for SSRIs in humans and it offers a plausible biological explanation for the delayed treatment response commonly observed in patients treated with SSRIs. While replication is warranted, these results have important implications for the design of future clinical studies investigating the neurobiological effects of SSRIs.


Radiosynthesis of [ 11 C]UCB-J
Proton irradiation of the target material (nitrogen-14 gas) was performed using the cyclotron: Scanditronix MC-32 with aluminium high-pressure gas target.Irradiations for carbon-11 were performed at 16 MeV.The target gas used was 10% hydrogen in nitrogen.Following irradiation, the target gas was transferred to the radiochemistry system (Scansys Aps) through stainless steel capillaries.[ 11 C]methyl iodide was synthesized from [ 11 C]methane by a standard circulation procedure.
Preparation of the precursor: The precursor (1.5 mg) was dissolved in MeOH (70 µL) and 1 M HCl (15 µL) was added.The resulting mixture reacted overnight at room temperature.On the day of synthesis, the liquid was removed by a stream of nitrogen to complete dryness.

Radiometabolite analysis
For measurement of intact tracer and radiolabeled metabolites, plasma samples up until 90 min were filtered through a 0.45 µm syringe filter (Whatman GD/X 13 mm, Cytiva, Buckinghamshire, UK) and subsequently diluted 1:1 with 20 mM disodium hydrogen phosphate and 5 mM sodium-1-decanesulfonate pH 7.2 with 2% isopropanol.Samples were analyzed in a fully automated column-switching HPLC system (UltiMate 3000, Thermo Fisher Scientific, Hvidovre, Denmark) connected to a radio-HPLC detector (PosiRam Model 2, LabLogic Systems, Sheffield, UK)(Gillings, 2009).The HPLC system was equipped with a small extraction column (Shimpack MAYI-ODS 30x4.6 mm, Shimadzu, Ballerup, Denmark) combined with an analytical column (Onyx Monolithic C18 50x4.6 mm, Phenomenex, Brønshøj, Denmark).For extraction and elution, the mobile phase consisted of 100% 20 mM disodium hydrogen phosphate and 5 mM sodium-1-decanesulfonate pH 7.2 with 2% isopropanol and 59% 100 mM sodium dihydrogen phosphate and 2 mM sodium-1-decanesulfonate pH 2.6 and 41% methanol, respectively.Samples were injected in a volume of 4 mL, and the analysis was run at a flow of 5 mL/min at 25 °C.The total runtime for each sample was 8.55 min with a 4 min extraction step, 4 min elution step and 0.55 min of equilibration.Four eluate fractions were collected in 2 min intervals, and radioactivity was subsequently measured using a gamma counter (Wizard 2480, Perkin Elmer, Wallac Oy, Turku, Finland).The parent tracer fraction was calculated as follows: % parent fraction = (radioactivity of parent eluate/total amount of collected radioactivity) x 100%.
Free fraction measurement [ 11 C]UCB-J free fraction was measured in the following way: plasma spiked with 1 µL/mL of tracer was added into one dialysis chamber (500 µL chambers, Harvard Apparatus, Holliston, MA, USA) and dialyzed against an equal volume of phosphatebuffered saline through a cellulose membrane (MWCO 10,000 Dalton, Harvard Apparatus, Holliston, USA).The system was incubated at 37 ºC for 30, 60, 120, 150 and 180 min.After completion of dialysis, samples were extracted and analyzed for radioactivity using the Cobra II gamma counter (Packard Instrument Company, Meriden, USA).The ratio of buffer:plasma radioactivity was plotted over time and fitted according to eq. 1 with a fixed average value of the diffusion coefficient (kD), using GraphPad Prism (v.9.0, GraphPad Software, San Diego, CA, USA) to determine fPeq.

Table S2 :
Effect of intervention groupGeneral linear model with age, sex and IQ as covariates to test group effect on [ 11 C]UCB-J VT * Intercept centered at age = 20 years and IQ = 100.

Table S3 :
Effect of intervention durationGeneral linear model with age, sex and IQ as covariates to test effect of intervention duration on [ 11 C]UCB-J VT *Intercept centered at age = 20 years, mean intervention duration (29 days), and IQ = 100.Rρ = partial correlation coefficient.

Table S7 :
Effects of intervention durationLikelihood-ratio tests of model with group-by-intervention duration vs. a nested model without group term.Age and sex included in both models