The most likely time for a woman to become depressed is after childbirth. ^{Reference Kendler, Neale, Kessler, Heath and Eaves1} Post-partum depression affects approximately 13% of women. ^{Reference O'Hara and Swain2,Reference García-Esteve, Ascaso, Ojuel and Navarro3} Post-partum depression has a great impact on the family and economy, and is considered a major public health problem. ^{Reference Petrou, Cooper, Murray and Davidson4,Reference Wisner, Chambers and Sit5} There is general agreement that the dramatic physiological changes that occur post-partum increase a woman's vulnerability to depressive symptoms, including post-partum depression. ^{Reference Bloch, Daly and Rubinow6,Reference Serreti, Olgiati and Colombo7}

Pregnancy and delivery are accompanied by hormonal changes as well as lower plasma tryptophan levels, both of which are thought to be aetiologically relevant to the mood changes that follow childbirth. ^{Reference Bloch, Daly and Rubinow6,Reference Maes, Ombelet, Verkerk, Bosmans and Scharpé8,Reference Payne9} Although plasma tryptophan availability is not directly related to mood changes, ^{Reference Maes, Ombelet, Verkerk, Bosmans and Scharpé8} the brain tryptophan availability index is decreased after delivery and is related to depressive symptoms. ^{Reference Baïlara, Henry, Lestage, Launay, Parrot, Swendsen, Sutter, Roux, Dallay and Demotes-Mainard10} The mood-lowering effects of experimental tryptophan depletion are controversial, ^{Reference Van der Does11} perhaps because of differences in 5-HTT genotype–tryptophan interaction. In women with previous depressive episodes, 5-HTT genotype may moderate the risk for depressive symptoms after tryptophan depletion. ^{Reference Moreno, Rowe, Kaiser, Chase, Michaels, Gelernter and Delgado12,Reference Neumeister, Hu, Luckenbaugh, Sachwrz, Nugent, Bonne, Herscovitch, Goldman, Drevets and Charney13} If childbirth is considered an environmental factor, there may be a strong pathophysiological link between post-partum mood changes and the genes that moderate 5-hydroxy-tryptamine (5-HT) signalling.

Jans et al ^{Reference Jans, Riedel, Markus and Blokland14} have proposed the concept of ‘serotonergic vulnerability’. Disruption of the serotonergic system may occur at several levels, including tryptophan availability, 5-HT synthesis, release, reuptake or metabolism, and/or pre- or post-synaptic 5-HT receptors. 5-HTT has received special attention because it plays a crucial role in the regulation of serotonergic function. ^{Reference Murphy, Li, Engel, Wichems, Andrews, Lesch and Uhl15,Reference Lesch and Gutknecht16}

In the light of these findings, we designed a prospective longitudinal multicentre study to evaluate interactions between 5-HTT genotype and post-partum mood changes. We hypothesised that 5-HTT genotype would shape the risk for depressive symptoms in post-partum women. We considered three genotype combinations of 5-HTTLPR and Stin2 VNTR polymorphisms that predict differential 5-HTT expression. ^{Reference Hranilovic, Stefulj, Schwab, Borrmann-Hassenbach, Albus, Jernej and Wildenauer17} Given that the acute decline in tryptophan availability that follows childbirth resembles experimental tryptophan depletion, we hypothesised that high-expression 5-HTT genotypes would be associated with depressive symptoms in the weeks following childbirth.

Method

Results

Participants

There were 1974 women who fitted the inclusion criteria for the study, 94 (5%) women refused to participate and 76 (3.8%) women were excluded because their EPDS questionnaires were incomplete. Thus, the final sample comprised 1804 women. At the 8-week follow-up, 1407 (78%) women remained in the study. At 32 weeks, 1337 (74.1%) women were evaluated (Fig. 1). Those women who dropped out during the follow-up period were compared with the final sample, revealing that women from lower social classes were the most likely to drop out (P=0.005). The mean age of the participants was 31.7 years (s.d.=4.6), range 18–46. Of the participants, 32% had attended primary school, 41% finished secondary school and 27% had a college degree. Most participants (68%) were employed, 9% were homemakers or students, 12% were unemployed and 11% were on sick leave or maternity leave. Forty-six per cent of the women were primiparous. Thirty-one per cent of the total sample has a family history of previous psychiatric treatment. Sixteen per cent had a previous personal history of psychiatric treatment.

Fig. 1 Follow-up study of post-partum depression; EPDS, Edinburgh Postpartum Depression Scale; MD, major depression: DSM–IV major depression episode; DIGS, Diagnostic Interview for Genetics Studies; PPD, post-partum depression.

Clinical variables

Of the 1407 women studied at 8 weeks post-partum, 214 scored 9/10 on the EPDS scale. A diagnosis of major depression was confirmed by DIGS in 112 (7.9%) women. At 32 weeks post-partum, 323 of 1337 women scored 9/10 on the EPDS, but only 61 (4.5%) new cases of major depression were confirmed by DIGS. Overall, 173 women (12.7%) had a major depression episode during the first 32 post-partum weeks (Fig. 1). There were no differences in socio-demographic variables (i.e. age, educational level, employment) between women diagnosed with major depression at 8 weeks post-partum and women diagnosed at 32 weeks post-partum.

Edinburgh Postnatal Depression Scale scores declined over the post-partum period. The mean EPDS score was 6.1 (s.d.=4.5) at baseline, 5.3 (s.d.=4.6) at 8 weeks post-partum and 4.4 (s.d.=4.7) at 32 weeks post-partum (Fig. 2).

Fig. 2 Edinburgh Postnatal Depression Scale (EPDS) score over time (2–3 days, 8 weeks and 32-weeks post-partum) in relation to the different 5-HTT genotype combinations. HE, no low-expressing genotype at either of the loci; ME, low-expressing genotype at one of the loci; LE, low-expressing genotypes at both loci.

Genotype frequencies according to serotonin transporter expression

No significant difference in the distribution of major depression according to genotype combination was observed at 8 weeks (P=0.089) or 32 weeks (P=0.125) post-partum.

At baseline, genotype was not a predicator of EPDS score. At 8 weeks after childbirth, EPDS scores were related to 5-HTT expression levels in a dose–response fashion (regression coefficient B=0.45, 95% CI 0.09 to 0.82, P=0.015). At 32 weeks post-partum, EPDS was not related to genotype (B=–0.14, 95% CI −0.53 to 0.26, P=0.49) (Table 1, Fig. 2). The test for interaction between time and expression level genotype suggested the difference between 8 and 32 weeks was not due to chance (χ²=2.7, d.f.=1, P=0.1).

Table 1 Frequency of different 5-HTT genotype combinations (5-HTT-GC) and Edinburgh Postnatal Depression Scale (EPDS) score at baseline (2–3 days post-partum), 8 weeks post-partum, and 32 weeks post-partum

	Baseline					8 weeks					32 weeks
Expression level genotypes	Mean	Regression coefficient (95% CI)	P	Mean	Regression coefficient (95% CI)	P	Mean	Regression coefficient (95% CI)	P
LE	5.85	0 a		4.89	0 a		4.31	0 a
ME	6.29	0.44 (–0.00 to 0.88)	0.052	5.49	0.37 (–0.10 to 0.84)	0.13	4.40	–0.28 (–0.79 to 0.23)	0.28
HE	6.08	0.23 (–0.54 to 1.00)	0.56	6.00	1.02 (0.18 to 1.88)	0.018	5.03	–0.08 (–0.98 to 0.83)	0.87
Regression coefficient linear trend b		0.24 (–0.10 to 0.58)	0.16		0.45 (0.09 to 0.82)	0.015		–0.14 (–0.53 to 0.26)	0.49

HE, no low-expressing genotype at either of the loci; ME, low-expressing genotype at one of the loci; LE, low-expressing genotypes at both loci

a. Reference category

b. Summary increase in EPDS score with one unit change in genotypic loading. P-values refer to the differences of ME and HE genotype with LE genotype as reference group

The differences remains significant after correcting for multiple comparison using Bonferroni correction between high expression (no low-expressing genotype at either of the loci) and low expression groups (low-expressing genotypes at both loci and low-expressing genotype at one of the loci) (P=0.045). We also introduce age in our linear regression model with not significant effect on the results (P=0.069).

Discussion

The incidence of post-partum depression in this study (12.7%) is in concordance with previous reports. ^{Reference O'Hara and Swain2,Reference García-Esteve, Ascaso, Ojuel and Navarro3} Although there was a trend (P=0.089), no significant interaction between the expression level genotype and major depression was found. The EPDS score at 8 weeks post-partum was associated with the high-expression genotype. These results are in agreement with our hypothesis that 5-HTT genotype may modulate the mood changes, mainly depressive symptoms, that women experience just after delivery.

A number of factors may explain why the 5-HTT genotype was significantly associated with the EPDS score at 8 weeks, but not with major depression. First, it could be that the lack of significance (P=0.089) may be a statistical problem related to the relatively small sample size of women with major depression (n=173) and/or a weak effect of the 5-HTT genotype. High-expression 5-HTT polymorphisms may promote tryptophan depletion and induce major depression post-partum, but only when other genetic and/or environmental factors are present (e.g. lack of social support or life events). It seems unlikely that EPDS score at 8 weeks may reflect the ‘blues’, a transient emotional liability that affects about 50% of post-partum women. ^{Reference Seyfried and Marcus27} The ‘blues’ are quantitatively and qualitatively different from depression; ^{Reference Miller, Rukstalis and Miller28} they emerge during the first post-partum week and disappear by the second week. Moreover, there is some evidence that the DSM–IV ²⁹ or ICD–10 ³⁰ definition of major depression is an arbitrary diagnostic convention imposed upon a continuum of depressive symptoms. ^{Reference Goldberg31–Reference Aggen, Neale and Kendler33}

Reduced availability of brain tryptophan during the post-partum period may explain the high (>9) EPDS scores. Experimental tryptophan depletion frequently leads to transient symptoms of depression in vulnerable individuals. ^{Reference Van der Does11} The brain tryptophan availability index decreases by 15% after delivery and is associated with depressive symptoms. ^{Reference Baïlara, Henry, Lestage, Launay, Parrot, Swendsen, Sutter, Roux, Dallay and Demotes-Mainard10} This reduction in the early post-partum period is not associated with plasma tryptophan levels ^{Reference Maes, Ombelet, Verkerk, Bosmans and Scharpé8} but rather with a dramatic increase in circulating levels of free amino acids that compete with tryptophan, such as leucine, isoleucine, valine and tyrosine, resulting in a significantly impaired transport of tryptophan across the blood barrier. The brain tryptophan availability index is calculated according to the Michaelis model for substrate competition on enzymes or transporters taking into account the total plasma tryptophan concentration and the blood concentration of competitor amino acids. So, the post-partum period is a ‘natural model’ of tryptophan depletion, and individuals with the LL 5-HTTLPR genotype indeed present with more depressive symptoms after tryptophan depletion. In a study of 43 individuals currently in remission from major depression, those with the LL genotype had significantly higher scores on the Hamilton Rating Scale for Depression (HRSD) than LS or SS carriers. ^{Reference Moreno, Rowe, Kaiser, Chase, Michaels, Gelernter and Delgado12} Moreno et al suggested that rapid uptake of 5-HT in people with LL, combined with decreased brain 5-HT availability during tryptophan depletion, produces a substantial decrease in serotonergic transmission, thereby enhancing depressive symptoms. ^{Reference Moreno, Rowe, Kaiser, Chase, Michaels, Gelernter and Delgado12} A recent study ^{Reference Neumeister, Hu, Luckenbaugh, Sachwrz, Nugent, Bonne, Herscovitch, Goldman, Drevets and Charney13} confirms that people with previous depressive episodes and the LL genotype have the greatest increase in HRDS scores after tryptophan depletion. The gene–tryptophan interaction is further supported by a double-blind placebo study of 15 healthy SS and 15 healthy LL volunteers. The SS genotype group outperformed the LL genotype group in tests of episodic memory and attention. ^{Reference Roiser, Müller, Clark and Sahakian34} Thus, it may be hypothesised that women with high-expression 5-HTT genotypes have an increased risk of depressive symptoms post-partum because serotonergic transmission is reduced. In line with the above observation, the genotype effect disappears by 32 weeks post-partum.

The decline of EPDS scores over the post-partum period is also consistent with previous observations. ^{Reference Evans, Heron, Francomb, Oke and Golding35,Reference Dennis36} Depressive symptoms in women with a high-expression 5-HTT genotype show the least change over time, possibly reflecting differences in the rate of brain tryptophan availability index normalisation during the post-partum period. Other aetiological mechanisms of post-partum depression, such as familiality, are in agreement with the time of onset observed in our study. ^{Reference Forty, Jones, Macgregor, Caesar, Cooper, Hough, Dean, Dave, Farmer, McGuffin, Brewster, Craddock and Jones37} In tryptophan depletion studies, high-expression 5-HTT genotypes are at risk, whereas low-expression variants have been implicated in life event interactions. ^{Reference Caspi, Sugden, Moffitt, Taylor, Craig, Harrington, McClay, Mill, Martin, Braithwaite and Poulton38–Reference Cervilla, Molina, Rivera, Torres-González, Bellón, Moreno, Luna, Lorente, Mayoral, King and Nazareth44} This suggests that gene effects on acute tryptophan depletion and stress response are distinct. According to the ‘serotonergic vulnerability’ model, ^{Reference Jans, Riedel, Markus and Blokland14} mood changes after delivery are the result of complex interactions between multiple variables, including acute hormonal changes and tryptophan depletion, which is mediated by 5-HTT genotype. In support of this hypothesis the risks of post-partum depression have been associated with a tryptophan hydroxylase gene polymorphism. ^{Reference Sun, Tsai, Ko, Chang and Yeh45}

To the best of our knowledge, there are two studies of 5-HTT expression during pregnancy and delivery. Coyle et al ⁴⁸ found a significant (P<0.003) over-representation of STin2.12 allele (the high expression allele) in 127 women with bipolar disorder, who later developed puerperal psychosis within a few days of delivery, compared with 380 controls. Scheid et al ^{Reference Scheid, Holzman, Jones, Friderici, Nummy, Symonds, Sikorskii, Regier and Fisher47} performed a cross-sectional study of 568 pregnant women to examine possible interactions between 5-HTTLPR polymorphisms and different types of life stressors relative to depressive symptoms. The only significant interaction identified was the sub-construct of ‘childhood abuse’. Their study, however, was conducted during pregnancy, not during the post-partum period, and therefore it is unlikely it tapped into specific mechanisms associated with tryptophan depletion.

The present study has several limitations. First, the brain tryptophan availability index was not measured. Second, the results pertain to White Spanish women and may or may not be applicable to other ethnic groups. Lastly, it is impossible to exclude the possibility that depressive symptoms account for the 27% attrition rate at 32 week post-partum. However, it is unlikely that genotype has a differential effect on attrition, making this an unlikely cause of bias. It is also worth mentioning several strengths of our study. First, our sample is relatively large. Second, the study was longitudinal and depression was evaluated categorically and dimensionally to increase sensitivity of the measure. Third, we analysed the combination of two (5-HTTLPR and Stin2 VNTR) polymorphisms related to functional expression. Finally, the homogeneity of our sample may be one of the principal methodological strengths. All participants were White Spanish women, all were young and, perhaps most importantly, all were evaluated during the very well-defined post-partum risk period.

In summary, our results suggest that there are no ‘bad’ or ‘good’ 5-HTT genotypes in relation to depression. High-expression 5-HTT genotypes might be a risk factor under certain environmental conditions such as tryptophan depletion after childbirth. This study supports a new hypothesis for understanding the biological mechanisms underlying depressive symptoms after delivery and encourages further study of gene–tryptophan interactions in mood disorders.