Pituitary volume mediates the relationship betweenpubertal timing and depressive symptoms duringadolescence

Sarah Whittle a,c, Murat Yucel a,c, Valentina Lorenzetti c, Michelle L. Byrne b,Julian G. Simmons a, Stephen J. Wood c,d, Christos Pantelis c,Nicholas B. Allen a,b,*

aOrygen Youth Health Research Centre, Centre for Youth Mental Health, University of Melbourne, AustraliabDepartment of Psychological Sciences, University of Melbourne, AustraliacMelbourne Neuropsychiatry Centre, Department of Psychiatry, University of Melbourne, Australiad School of Psychology, University of Birmingham, UK

Received 14 April 2011; received in revised form 18 October 2011; accepted 18 October 2011

Psychoneuroendocrinology (2012) 37, 881—891

KEYWORDSPituitary;Depression;Adolescence;Puberty;Pubertal timing;Risk factors

Summary Early timing of puberty (i.e., advanced pubertal maturation relative to peers) hasbeen linked to the onset of depressive symptoms during the early adolescent phase. However, theprecise neurobiological mechanisms linking early pubertal timing to adolescent depressive symp-toms are not clear. We investigated whether the volume of the pituitary gland, a key component ofthe hypothalamic—pituitary—gonadal (HPG) and hypothalamic—pituitary—adrenal (HPA) axes,mediated the relationship between pubertal timing and depressive symptoms in 155 adolescents(72 females) both cross-sectionally and longitudinally. At baseline (M age 12.7, SD 0.5 years), earlypubertal timing predicted larger pituitary gland volume and higher depressive symptoms (especiallyfor girls), but there was no mediation effect. Longitudinally, however, larger pituitary gland volumeat baseline was found to mediate the relationship between early pubertal timing and increaseddepressive symptoms over time (M follow-up period = 2.57 years, SD = 0.26) for both boys and girls.Our findings suggest that neurobiological mechanisms are partly responsible for the link betweenearly pubertal timing and depressive symptoms in adolescents. We speculate that an enlargedpituitary gland in adolescents with early pubertal timing might be associated with hyperactivationof the hormonal stress response, leading to increased susceptibility to environmental stressors, andsubsequent development of depressive symptoms. Given the well-established relationship betweenincreasing depressive symptoms in adolescence and later disorder, these findings have implicationsfor targeted prevention and early intervention strategies for depressive disorders in adolescence.# 2011 Elsevier Ltd. All rights reserved.

* Corresponding author at: Orygen Youth Health Research Centre and Department of Psychological Sciences, University of Melbourne, Victoria3010, Australia. Tel.: +61 3 9342 2819; fax: +61 3 9342 2941.

E-mail address: nba@unimelb.edu.au (N.B. Allen).

Available online at www.sciencedirect.com

j our na l h omepa g e: www.e l se v ie r.c om/l oca te/ psyne ue n

0306-4530/$ — see front matter # 2011 Elsevier Ltd. All rights reserved.

doi:10.1016/j.psyneuen.2011.10.004

882 S. Whittle et al.

1. Introduction

The incidence of depressive symptoms rises sharply duringthe early to mid adolescent phase (Lewinsohn et al., 1993,1998), and there is a well-established relationship betweenincreasing depressive symptoms during this period and riskfor later depressive disorder (Shankman et al., 2009). While anumber of factors have been proposed to contribute to riskfor depression during adolescence (e.g., family history,adverse environments, cognitive style, etc. Weissmanet al., 1992; Hammen, 1995; Alloy et al., 2001), issues thatrelate to pubertal development have also been suggested ascritical (Angold et al., 1998; Angold and Costello, 2006).

Puberty involves major hormonal changes that occur as aresult of activation of the hypothalamic—pituitary—gonadal(HPG) axis, which culminates in gonadal maturation. Thesebiological events manifest in secondary sex characteristics(e.g., menarche and morphologic development) (Sisk andFoster, 2004). Pubertal development, measured either viagonadal hormones or gonadotropins, or via body morphologic(i.e., Tanner) stage, have all been associated with higher riskfor depression, with some evidence to suggest that thisrelationship is stronger for girls (Susman et al., 1985; Angoldand Costello, 2006; Patton et al., 2008). However, findingshave not been entirely consistent (see Angold and Costello,2006). In particular, the timing of puberty (i.e., the timing ofchanges in pubertal status relative to peers), rather thanpubertal stage at a given time-point, has been suggested tobe most important in predicting adolescent depression(Angold and Costello, 2006). Early puberty has been foundto predict depressive symptoms and disorders in girls in manystudies (Ge et al., 2001a; Kaltiala-Heino et al., 2003; e.g.,Angold and Costello, 2006), although there are some contra-dictory findings (Graber et al., 1997). The literature for boysis more mixed, with some studies finding that earlier, andsome finding that later onset of puberty (and some findingthat both early and late onset) is associated with depressivesymptoms and disorders (Graber et al., 1997; Kaltiala-Heinoet al., 2003; Angold and Costello, 2006; Conley and Rudolph,2009).

Notwithstanding some inconsistencies in the literature,findings to date clearly demonstrate a link between pubertaltiming and depression. However, the mechanisms underlyingthis link are not clear. It has been variously suggested thatbiological, social and psychological processes play a role(Kaltiala-Heino et al., 2003), but there has been little empiri-cal investigation into these possibilities. A few studies haveinvestigated potential psychological mediators between pub-ertal timing and depression (e.g., Graber et al., 2006), andone study (Natsuaki et al., 2009) has investigated cortisolreactivity to interpersonal stressors as a mediator betweenpubertal timing and internalizing symptoms. The findingsdemonstrated that the higher levels of internalizing pro-blems seen in early maturing girls were at least partiallyattributable to their heightened cortisol reactivity to inter-personal stress.

To date, no study has investigated neurobiological med-iators of the link between pubertal timing and depression.Among potential neurobiological mediators, structural prop-erties of the pituitary gland might be particularly significant.The pituitary gland is an integral part of the HPG axis, whichbecomes activated at puberty and underlies development of

reproductive maturity. Gonadal releasing hormone secretionat puberty travels to the pituitary to signal the synthesis andsecretion of the pituitary gonadotropins (luteinizing hor-mone, LH, and follicle stimulating hormone, FSH), whichact on target cells in the testes and ovaries to direct theproduction of sperm and eggs, as well as the secretion ofsteroid hormones. There is evidence that the size of thepituitary gland may be used to reflect HPG activity (Peperet al., 2010). Indeed, animal and human (i.e., adolescentsamples) studies have shown high levels of sex steroids to beassociated with an increased cell proliferation (Nolan andLevy, 2006a,b) and volume (Peper et al., 2010) of the pitui-tary gland. Human developmental studies have shown thatthe size of the pituitary gland increases during adolescence,with the increase appearing earlier for girls (i.e., mirroringthe earlier onset of puberty in girls, Elster et al., 1990). Wehave recently shown that adolescent pubertal status is sig-nificantly positively correlated with pituitary volumes(Zipursky et al., in press).

There is also some evidence that size of the pituitary glandis associated with pubertal timing and depression vulnerabil-ity. For example, Kao et al. (1992) found that children withcentral precocious puberty (premature reactivation of theHPG axis) had larger pituitary gland volumes than age-matched controls. While adult studies have yielded incon-sistent results regarding changes in pituitary volume indepression (Krishnan et al., 1991; Lorenzetti et al., 2009),adolescent studies are consistent in finding enlarged pitui-tary volumes to be associated with depression both prospec-tively and longitudinally (MacMaster and Kusumakar, 2004;MacMaster et al., 2006; Zipursky et al., in press).

Given these relationships between pubertal timing andpituitary volume, and pituitary volume and depression, thecurrent study aimed to investigate whether pituitary volumemight mediate the link between early pubertal timing anddepressive symptoms in adolescence. We hypothesized thatearlier pubertal timing would be associated with increaseddepressive symptoms (especially in girls) and that larger sizeof the pituitary volume would mediate this link. In a cohort of155 adolescents, we employed a longitudinal design wherebywe assessed depressive symptoms at two time points (earlyand mid adolescence) allowing us to investigate mediatingeffects both cross-sectionally and prospectively. This studyextended our previous work showing a link between pituitaryvolume and the development of internalizing symptoms dur-ing adolescence (Zipursky et al., 2011) in two importantways. Firstly, we introduced pubertal timing as a potentiallyimportant predictor of depressive symptom developmentduring adolescence, and secondly, we investigated the roleof pituitary gland volume as a mediator (i.e., mechanism) ofthe link between pubertal timing and depressive symptomdevelopment.

2. Methods

2.1. Participants

Participants completing baseline assessments (i.e., ques-tionnaires and Magnetic Resonance Imaging [MRI] scans)were 155 adolescents (83 males, M age 12.7, SD 0.5 years;72 females, M age 12.6, SD 0.4 years) recruited from schoolsacross metropolitan Melbourne, Australia. At baseline, no

Pituitary volume mediates the link between pubertal timing and depression in adolescence 883

participant met DSM-IV criteria for a depressive disorder(see Yap et al., 2008 and Supplement 1 for further details).One hundred and forty-one participants (90%) from thissample (65 females) completed a follow-up assessment(diagnostic interview and questionnaires) approximately2—3 years after the baseline assessment (i.e., M follow-upperiod = 2.57 years, SD = 0.26). Informed consent wasobtained from all participants (i.e., adolescent and parent),in accordance with the guidelines of the Human ResearchEthics Committee of the University of Melbourne, Australia.

2.2. Neuroimaging

2.2.1. Image acquisition and analysisAt baseline, MRI scans were performed on a 3 T GE scanner,using a gradient echo volumetric acquisition sequence (i.e.,repetition time = 36 ms; echo time = 9 ms; flip angle = 358,field-of-view = 20 cm2, matrix = 410 � 410) to obtain 124 T1-weighted contiguous slices (i.e., voxel dimensions =0.4883 mm � 0.4883 mm � 1.5 mm).

Tracing was performed by a trained research assistant oneach individual’s images using the software package ANALYZE(Mayo Clinic, Rochester, USA; http://mayoresearch.-mayo.edu/mayo/research/robb_lab/). The pituitary wasdefined and quantified based on a previously published tech-nique (Pariante et al., 2004). Specifically, images were tracedin the coronal plane, where the pituitary is best visualized(Pariante et al., 2004, see Fig. 1; Lorenzetti et al., 2009). Weexcluded the infindibular stalk from the tracing, but includedthe hyper-intense region in the posterior pituitary, which isthought to represent high levels of vasopressin concentrations(Sassi et al., 2001). The boundaries of the pituitary wereclearly defined by the diaphragma sellae, superiorly; thesphenoid sinus, inferiorly, and the cavernous sinuses, bilater-ally (Garner, 2004; Pariante et al., 2004; Garner et al., 2005).Pituitary volume estimates were calculated by summing allvoxels within the traced region on each slice. Intraclass cor-relation coefficients (absolute agreement) for intra and inter-rater reliabilities, assessed on ten randomly selected images,were 0.911 and 0.950, respectively.

Intracranial volumes (ICV) were delineated from a sagittalreformat of the original 3D-dataset. The major anatomicalboundary was the dura mater below the inner table andgenerally visible as a white line. Where the dura materwas not visible, the cerebral contour was outlined. Other

Figure 1 Traced pituitary gland on coronal slice of MR image.

landmarks included the undersurfaces of the frontal lobes,the dorsum sellae, the clivus, and the posterior arch of thecraniovertebral junction (Eritaia et al., 2000).

2.3. Psychiatric symptoms

At baseline and follow-up, adolescents completed an assess-ment of self-reported depressive symptoms using the Centerfor Epidemiological Studies-Depression Scale (CES-D, Radl-off, 1977). The CES-D is a self-report questionnaire designedto measure depressive symptoms in the general population.The questionnaire contains 20 items that relate to mood,somatic complaints, relations with others, and motor func-tioning during the past week. The CES-D has been found to bea valid and reliable measure for use in adolescent populations(Garrison et al., 1991).

2.4. Pubertal status

At baseline and follow-up, one parent completed the Pub-ertal Development Scale (PDS) on behalf of their child. Atfollow-up, the child also completed the PDS. For girls, thismeasure includes 12 items assessing the stage of breastdevelopment, hair growth, acne presence, hip width andmenarcheal status. For boys, this measure includes 11 itemsassessing genitalia development, hair growth, acne presenceand voice change. Reliability and validity of the PDS has beenwell established (Brooks-Gunn et al., 1987).

At follow-up, the child rated their current developmentwith the aid of schematic drawings of secondary sexualcharacteristics based on the five Tanner stages (Marshalland Tanner, 1969; Morris and Udry, 1980). Girls rated them-selves on breast development and pubic hair growth, andboys on their genitalia development. Such ratings have beenshown to correlate well with physician examinations (Morrisand Udry, 1980; Dorn et al., 1990; Schlossberger et al., 1992).

2.5. Attrition analysis and missing data

There were no differences between those participants whodid and did not complete the follow-up assessment in termsof pituitary volumes (t(1,150) = 0.18, p = 0.86), baselinedepressive symptoms (t[1,141] = 1.07, p = 0.28), or baselineparent-reported pubertal status (t(1,142) = 0.46, p = 0.64).Of the 155 participants that completed baseline assessments,CES-D scores were not available for 12 participants andparent-reported pubertal status was not available for afurther 11 participants. Of the 141 participants that com-pleted follow-up assessments, CES-D scores were not avail-able for 3 participants, parent-reported pubertal status wasnot available for a further 6 participants and child-reportedpubertal status was not available for 22 participants. Pitui-tary volumes were not available for 2 participants due toexcessive artefact in their T1-weighted images. Missingvalues (i.e., for both baseline and follow-up measures) weretreated as occurring at random, and were estimated based onmaximum-likelihood procedures that preserve the distribu-tional characteristics of the non-missing values on the itemand that takes into account the relevant longitudinal designcharacteristics (Schafer and Graham, 2002) and the long-itudinal structures in the data (Schafer, 2001).

Table 1 Descriptive statistics for all variables and tests for sex differences.

n (Males/Females)

Males Females t

M (SD) Range M (SD) Range

Age at T1 82/72 12.64 (0.48) 11.4—13.6 12.58 (0.41) 11.5—13.7 0.83Age at T2 76/65 15.04 (0.49 13.7—16.1 15.00 (0.41) 13.9—16.0 0.63CES-D at T1 77/65 13.22 (10.08) 0—51 10.58 (9.15) 0—55 1.62CES-D at T2 74/64 8.81 (6.84) 0—37 11.53 (9.35) 0—43 1.93Pituitary volume (mm3) 80/71 437.2 (136.3) 120—915 506.1 (138.8) 202—879 3.07 **

ICV (mm3) 82/71 1584364.9(119,804)

1,264,110—1,958,200 1460322.7(108805.2)

1,211,010—176,6760 6.66 ***

T1 PR score 75/68 14.41 (4.83) 9—32 16.04 (4.94) 8—34 2.00 *

T2 PR score 75/60 21.15 (6.64) 6—38 21.33 (5.36) 5—34 0.18T2 SR score 65/54 25.89 (5.60) 12—38 22.20 (4.04) 15—32 4.17 ***

Male Tanner 64 4.05 (0.68) 2—5Female Tanner (Br) 50 4.14 (0.73) 2—5Female Tanner (Pu) 50 4.10 (0.71) 2—5

Note. CES-D, Center for Epidemiological Studies-Depression Scale; PR, parent-report; SR, self-report; Br, breast development; Pu, pubic hairgrowth; T1, baseline; T2, follow-up.* p < 0.05.** p < 0.01.*** p < 0.001.

884 S. Whittle et al.

2.6. Data analysis

Descriptive statistics (and n’s) for all variables are shown inTable 1. The distribution of pituitary gland volume in oursample compares well with other available data using sam-ples with similar ages (e.g., MacMaster et al., 2007; Peperet al., 2010). Pituitary volumes were positively skewed andwere square root transformed in order to ensure a normaldistribution of values.

2.6.1. Creation of pubertal timing measurePrior to data imputation, a pubertal timing measure wascreated separately for each sex using the parent- andchild-report PDS scores and the child-report Tanner linedrawings; the rationale for this being that a single measureon its own would be affected by specific methodologicalartefacts (i.e., self-report bias or lack of parental insight).Further, all measures were meaningfully correlated (seeTables 2 and 3). For girls, 90% of correlations betweenmeasures were significant, and the single non-significantcorrelation was in the expected direction. For boys, all

Table 2 Correlations among female pubertal timing mea-sures at T1 and T2.

1 2 3 4 5

1.T1 PR score — 0.50 ** 0.48 ** 0.27 * 0.31 **

2.T2 PR score — 0.42 ** 0.21 0.25 *

3.T2 SR score — 0.54 ** 0.44 *

4.Female Tanner (Br) — 0.48 **

5.Female Tanner (Pu) —

Note. PR, parent-report; SR, self-report; Br, breast development;Pu, pubic hair growth; T1, baseline; T2, follow-up.* p < .05.** p < .01.

correlations between measures were significant and effectsizes were moderate to high.

To create the pubertal timing measure, all measures ofpubertal status were adjusted for age (separately for eachsex) and entered into an exploratory factor analysis (EFA)using Maximum Likelihood extraction. For girls, one factoremerged with an eigenvalue of greater than 1. This factoraccounted for 52.8% of the variance in measures and load-ings ranged from 0.49 to 0.79. For boys, one factor emergedwith an eigenvalue of greater than 1. This factor accountedfor 61.6% of the variance in measures and loadings rangedfrom 0.53 to 0.85. A pubertal timing score was then gen-erated for girls and boys as the standardized factor scorederived from the EFAs described above. Higher scoresrepresent more advanced pubertal development (i.e., ear-lier timing relative to peers of the same age). This con-ceptualization and operationalization of pubertal timing(i.e. level of development relative to age) is consistentwith past and recent theory and research on pubertal timing(Steinberg, 1987; Ge et al., 2001b; Dorn et al., 2003;Rudolph and Troop-Gordon, 2010).

Table 3 Correlations among male pubertal timing measuresat T1 and T2.

1 2 3 4

1.T1 PR score — 0.61 ** 0.36 ** 0.25 *

2.T2 PR score — 0.61 ** 0.49 **

3.T2 SR score — 0.63 **

4. Male Tanner —

Note. PR, parent report; SR, self-report; T1, baseline; T2, follow-up.* p < 0.05.** p < 0.01.

Pituitary volume mediates the link between pubertal timing and depression in adolescence 885

2.7. Mediation analysis

Two sets of mediation analyses were performed. For thefirst (cross-sectional) set of analyses, age-corrected base-line CES-D score (i.e., residual score from regressing CES-Dscore on age) was the outcome variable, and for the second(longitudinal) set of analyses, change in CES-D score frombaseline to follow-up was the outcome variable (i.e., resi-dual score from regressing follow-up CES-D score on base-line CES-D score). For all analyses, transformed pituitaryvolumes were corrected for age and ICV (using linearregression, separately by sex, Free et al., 1995) prior toentry into mediation models. All analyses were performedusing macros developed by Preacher and Hayes (2004)implemented in PASW 18. Mediation was tested by assessingthe significance of the cross product of the coefficients forthe IV (pubertal timing) to mediator (pituitary volume)relation (the a path), and the mediator to DV (change indepressive symptoms) relation controlling for the IV (the bpath). The a-b cross product tests the statistical signifi-cance of the difference between the total effect, or c path,and the direct effect, or c0 path, which is the impact of theIV on the DV adjusting for the effect of the mediator. Sexwas used as a covariate in the model. In a second analysis,sex was tested as a moderator of the mediation effect. In allanalyses, a bootstrapping method was applied to test thesignificance of the mediation effect (Preacher and Hayes,2004). Rather than impose questionable distributionalassumptions, bootstrapping generates an empirical approx-imation of the sampling distribution of a statistic byrepeated random resampling from the available data,and uses this distribution to calculate p-values and con-struct bias corrected and accelerated confidence intervals(for these analyses, 3000 resamples were taken and 95%confidence intervals were used) (see Hayes, 2009b, fordetails). If the confidence intervals do not contain zero,the point estimate is significant at the level indicated.

This method provides advantages to Baron and Kenny’s(1986) approach, which fails to provide a specific statisticaltest for the indirect effect that an independent variable hason a dependent variable via a proposed mediator. Further,using this method enabled us to test for significant mediationin the absence of a direct effect of the IV on the DV. That theIV can exert an indirect effect on the DV through the med-iator in the absence of an association between the IV and DVhas been argued as a possible, if not common, scenario(MacKinnon et al., 2000; Hayes, 2009a) and is viable becausea total effect is the sum of many different paths of influence,direct and indirect, not all of which may be a part of theformal model. For example, it could be that two or moreindirect paths carry the effect from X through Y, and thosepaths operate in opposite directions. These indirect effectswith opposite signs can cancel each other out, producing atotal effect that is not detectably different from zero, inspite of the existence of specific indirect effects that arenot zero.

3. Results

Scatterplots showing relationships between all variables arepresented in Fig. 2.

3.1. Cross-sectional analyses

As can be seen in Fig. 3, there was a significant associationbetween pubertal timing and baseline depressive symptoms(path c), with earlier pubertal timing predicting highersymptoms. Earlier pubertal timing also predicted largerpituitary volumes (path a). However, pituitary volume wasnot significantly associated with depressive symptoms (pathb), nor was there significant mediation (i.e., the a—b cross-product was not significant as indicated by the 95% confi-dence interval containing zero).

Because moderated mediation is possible in the absenceof simple mediation (Preacher and Hayes, 2004), we testedthe moderating role of sex in the above analysis. Althoughthere was no significant moderated mediation (neither theinteraction between pubertal timing and sex in the mediatormodel [B = 10.59, SE = 26.67, t = 0.40, p = 0.692] nor theinteraction between pituitary volume and sex in the depen-dent variable model [B = 0.02, SE = 0.01, t = 1.37, p = 0.174]were significant using model 5 in the macro), there was asignificant sex difference in the association between pubertaltiming and baseline symptoms (B = �6.46, SE = 2.0,t = �3.23, p = 0.002). Running analyses separately by sexrevealed that earlier pubertal timing predicted higher symp-toms for girls (B = 6.14, SE = 1.42, t = 4.33, p < 0.001) but notboys (B = �0.32, SE = 1.32, t = �0.24, p = 0.811).

3.2. Longitudinal analyses

In accordance with the cross-sectional analyses above, long-itudinal analyses showed that earlier pubertal timing wassignificantly associated with larger pituitary volume (seeFig. 4, path a) and larger pituitary volume was associatedwith higher follow-up depressive symptoms (controlling forbaseline symptoms). Neither the total effect of pubertaltiming on follow-up depressive symptoms (path c, i.e., notcontrolling for pituitary volume), nor the direct effect (pathc0, i.e., controlling for pituitary volume) was significant.However, bias-corrected 95% confidence intervals showedthat larger pituitary volume significantly mediated the rela-tionship between earlier pubertal timing and higher follow-up depressive symptoms (CI: 0.09—1.12, M = 0.01, SE = .25).

In the subsequent moderated mediation analysis, wefound that sex did not significantly moderate the indirecteffect of pubertal timing on change in depressive symptomsthrough pituitary volume (neither the interaction betweenpubertal timing and sex in the mediator model [B = 10.59,SE = 26.67, t = 0.40, p = 0.692], nor the interaction betweenpituitary volume and sex in the dependent variable model[B = 0.00, SE = 0.01, t = 0.26, p = 0.794] was significant usingmodel 5 in the macro). Although there was no significantmoderated mediation, there was a main effect of sex onfollow-up symptoms (B = �2.71, SE = 1.14, t = �2.38,p = 0.018), with girls having higher symptoms than boys.

4. Discussion

The precise neurobiological mechanisms linking early pub-ertal timing to depressive symptoms are currently unclear.The aim of this study was to investigate whether pituitaryvolume mediated the relationship between pubertal timing

Figure 2 Scatterplots showing relationships between all raw variables. R2 values are given to indicate strength of relationshipsseparately for males and females. Note that the y-axis for panels (b) and (d) represents the unstandardized residuals from regressingfollow-up CES-D scores onto baseline CES-D scores. The x-axis for panels (c), (d) and (e) represent factor scores; higher values indicateearlier pubertal timing.

886 S. Whittle et al.

Figure 3 Cross-sectional mediation analysis (N = 155). Path a is the total effect of pubertal timing on pituitary volume, path b is thetotal effect of pituitary volume on baseline depressive symptoms, path c is the total effect of pubertal timing on baseline depressivesymptoms (i.e., not controlling for pituitary volume), and path c0 is the direct effect of pubertal timing on baseline depressivesymptoms (i.e., controlling for pituitary volume). T1 = Time 1 (baseline).

Pituitary volume mediates the link between pubertal timing and depression in adolescence 887

and depressive symptoms in a large sample of adolescentsusing a longitudinal design. Our results did support the role ofthe pituitary gland in mediating the link between earlypubertal timing and increasing depressive symptoms fromearly to mid adolescence. Interestingly, we did not find such amediation effect cross-sectionally (during early adoles-cence). Further, while sex did not moderate the longitudinalmediation effect, there were sex differences in the directrelationship between pubertal timing and depressive symp-toms, whereby only girls with earlier pubertal timing experi-enced increased symptoms.

Our results provide evidence that neurobiological factorsmay partly underlie the commonly reported link betweenearly pubertal timing and depressive symptoms in adoles-cence. In particular, a larger size of the pituitary glandappeared to mediate this relationship. That we found earlierpubertal timing in adolescents to be associated with largervolume of the pituitary gland is consistent with evidence thatthe onset of puberty is associated with activation of the HPGaxis, which leads to increased cell proliferation in the pitui-tary gland as a result of the release of sex hormones triggeredby the hypothalamus (Peper et al., 2010). This finding is alsoconsistent with a study showing that children with centralprecocious puberty (early pubertal onset caused by early HPG

Figure 4 Longitudinal mediation analysis (N = 155). Path a is the ttotal effect of pituitary volume on follow-up depressive symptoms, pasymptoms (i.e., not controlling for pituitary volume), and path c0

symptoms (i.e., controlling for pituitary volume). T2 = Time 2 (followin this analysis, so that the outcome variable represents change in

axis activation) had increased pituitary volume compared toage-matched controls (Kao et al., 1992).

The mechanisms by which enlarged pituitary volumemight then lead to increased depressive symptoms areunclear, but it is reasonable to speculate that the role ofthe pituitary gland in hypothalamus—pituitary—adrenal(HPA) axis activation may be important. Indeed, while thepituitary gland is a part of the HPG axis and plays a role insexual development, it is also an integral part of the HPA axis,whose activity mediates, by a negative feedback loop, cir-culating levels of stress hormones (i.e., cortisol). Severalresearchers have shown that the dramatic alterations in sexsteroids seen with the onset of puberty play a potentialmodulatory role in HPA axis function, possibly contributingto vulnerability for depression in adolescents (e.g., Youngand Altemus, 2004; Stroud et al., 2006). Further, a largerpituitary volume is thought to be associated with relativelyhigher activity of the HPA axis (Westlund et al., 1985; Gertzet al., 1987; Axelson et al., 1992). Thus, we suggest that theprospective relationship between increased pituitary volumeand increasing depressive symptoms might reflect hyperac-tivation of the hormonal stress response, leading to increasedsusceptibility to environmental stressors. This is consistentwith the work of Natsuaki et al. (2009), who found that

otal effect of pubertal timing on pituitary volume, path b is theth c is the total effect of pubertal timing on follow-up depressiveis the direct effect of pubertal timing on baseline depressive-up). Note that baseline depressive symptoms are controlled fordepressive symptoms over time.

888 S. Whittle et al.

increased cortisol response to interpersonal stressorsmediated the relationship between earlier pubertal timingand internalizing problems in girls. It is also consistent withwork suggesting that the link between early puberty anddepression in girls is associated with an increased vulner-ability to stressful life events (Ge et al., 2001a). Furthersupport for this notion comes from previous studies that haveshown the pituitary to be larger in depressed patients whoshow less suppression of cortisol secretion by dexamethasoneand thus have a more hyperactive HPA axis (Krishnan et al.,1991; Axelson et al., 1992). Interestingly, HPA axis hyper-activity in depressed patients has been interpreted as show-ing a lack of negative inhibitory feedback by circulatingglucocorticoid hormones on the HPA axis, especially at thelevel of the pituitary (Pariante et al., 2002; Pariante, 2003).It is also possible that this HPA-axis hyperactivity might leadto depressive symptoms in adolescence due to a disruption orfailure of HPA axis control by prefrontal regions (Sullivan andGratton, 2002; Herman et al., 2003), which are undergoingmarked developmental changes over adolescence (reviewedin Spear, 2000) and hence have not reached mature levels offunction.

Interestingly, pituitary volume was found to mediate therelationship between pubertal timing and depressive symp-toms only when considering change in depressive symptomsover time (i.e., there was no mediation cross-sectionally fordepressive symptoms during early adolescence). During earlyadolescence, although there was no mediating effect ofpituitary volume, there was a direct effect of early pubertyon concurrent high depressive symptoms. As such, it ispossible that mediating factors other than pituitary glandvolume might play a stronger role, such as psychosocialfactors including increased stress or poor coping associatedwith earlier physical signs of maturity (Brooksgunn et al.,1985). While such psychosocial factors are also likely to beassociated with biological changes (which may have fasteracting depressogenic effects), it is possible that only overtime might the consequences of an enlarged pituitary volume(i.e., increased depressive symptoms) become evident. Forexample, it has been suggested that stress at key periods ofsynaptic organization could modify the trajectories of neuralconnections, leading to an incubation period, such that theeffects of stress are not apparent at the time of adversity butmay emerge later, when the synaptic organization has beencompleted (Lupien et al., 2009). Thus, given the markedbrain reorganization (particularly of the prefrontal cortex)occurring during adolescence, and the role of the prefrontalcortex in HPA-axis regulation, it is conceivable that in sus-ceptible individuals (i.e., those with enlarged pituitaryvolumes), exposure to stressful experiences during earlyadolescence (a period of neural vulnerability) sets intomotion a series of events that lead to a heterotypic reorga-nization of synaptic development, resulting in a protractedexpression of depression.

During early adolescence, we found that the direct effectof earlier pubertal timing on increased depressive symptomswas significant only for girls. This finding is consistent withmuch evidence showing that early puberty is associated witha range of maladaptive outcomes in girls, particularly depres-sive symptoms and disorders (see Kaltiala-Heino et al., 2003;Angold and Costello, 2006). In boys, the literature has beenmore mixed, and our results are consistent with others

finding no effect of pubertal timing on depression (seeKaltiala-Heino et al., 2003; Angold and Costello, 2006). Thatthere were no sex differences in the mediated or directeffects of pubertal timing on follow-up symptoms is surpris-ing, given our finding of sex differences in pituitary volume,and other longitudinal research showing sex differences inthe effect of pubertal timing on depression in later adoles-cence/young adulthood (Graber et al., 2004). Firstly, it maybe that pituitary volume (at least in early adolescence) is notan early precursor underlying later sex differences in depres-sion. Secondly, it may be that sex differences in relationshipsmay emerge later in adolescence, when sex differences indepression (females > males) become more pronounced.Indeed, in our sample, there were no sex differences indepressive symptoms at baseline or follow-up.

4.1. Strengths and limitations

Strengths of this study include the longitudinal design and themulti-informant measures of pubertal status. A number oflimitations, however, must be acknowledged. First, while wehave claimed to be assessing pubertal timing, it is difficult toseparate pubertal timing from pubertal stage. That is, in asample of adolescents who have not yet completed pubertaldevelopment, those with earlier timing are also likely to be ata later stage of development. It is only after puberty, whenall adolescents are fully mature, that pubertal timing isunambiguously unconfounded with stage (Angold and Cost-ello, 2006). Second, although we collected both parent andchild reports of pubertal stage, limiting potential biases thatmight be introduced from collecting a single measure, we didnot collect more objective measures of pubertal stage suchas physical examination by a qualified clinician, or hormonalassays that measure adrenal and gonadal (or adrenal/gona-dal-releasing) hormones upstream from their external phy-sical effects. Collecting a wider range of measures wouldallow investigation of the timing of different aspects ofpubertal development (Blakemore et al., 2010). Third, it ispossible that there may have been a testing bias with regardto our longitudinal assessment of depressive symptoms. Inour sample, female depressive symptoms (on average)remained fairly constant over time, and male symptomsdecreased. Although this pattern is not inconsistent withsome previous research (Angold et al., 1996; Tram and Cole,2006; Montague et al., 2008), it is in contrast to muchresearch showing increases in depressive symptoms overadolescence, particularly for girls. It has been suggestedthat testing effects (i.e., effects of successive administra-tions of a questionnaire) may be partly responsible fordecreases in scores over time in longitudinal studies (Twengeand Nolen-Hoeksema, 2002). Fourth, our suggestion regard-ing the role of stress and the HPA axis in the link betweenpituitary volume and subsequent increasing depressive symp-toms is speculative. Future work would benefit from collect-ing measures of stress and HPA axis function to investigatethese possibilities. Further, we cannot be certain about thedirection of effects in our findings. Although not part of ourtheoretical model, it is possible that pituitary volume mayinfluence the timing of puberty. Future work assessing pitui-tary volume during childhood, before the onset of puberty,would help to tease apart this possibility. Fifth, althoughtrajectories of depressive symptom change during this age

Pituitary volume mediates the link between pubertal timing and depression in adolescence 889

period have been found to be clinically significant (Dekkeret al., 2007), future work should investigate whether thepresent findings hold for the prediction of case-level depres-sive disorder. Sixth, the specificity of our finding is unknown.It may be that enlarged pituitary volume during adolescenceis a risk factor for the development of a range of psychiatricconditions. Other work has shown, for example, thatenlarged pituitary volume during adolescence represents arisk marker for future development of psychosis (Garneret al., 2005). However, this research was performed in asample of older adolescents. Given the marked growth of thepituitary gland over adolescence, its volume at any one timepoint may have specific influences on mental health out-comes depending on the developmental and social context.

5. Conclusions

In this study we found evidence that the volume of the pituitarygland mediates the relationship between early pubertal timingand increasing depressive symptoms across early to mid-ado-lescence. This finding suggests that neurobiological mechan-isms are partly responsible for the link between early pubertaltiming and adverse mental health outcomes in adolescents; inparticular, we speculate that an enlarged pituitary glandvolume in adolescents with early pubertal onset might beassociated with hyperactivation of the hormonal stressresponse, leading to increased susceptibility to environmentalstressors, and subsequent development of depressive symp-toms. Given that adolescents with increased depressive symp-toms are at increased risk of developing depressive disordersduring later adolescence and adulthood, these findings mayhave implications for prevention and early intervention stra-tegies for depressive disorders in adolescence. For example,our findings suggest the potential utility of preventativeefforts targeting early maturing girls and boys. They alsosuggest that treatment or prevention efforts in early maturingadolescents should take into account a biological componentto aetiology by recognizing that the stress system might bebiologically hyperactive in these individuals, and as suchinterventions that seek to modulate emotional reactions oncethey are elicited might be less successful than those that seekto prevent or reduce environmental exposure to stressorsamong these individuals.

Role of funding source

This research was supported by grants from the ColonialFoundation, the National Health and Medical Research Coun-cil (NHMRC; Australia; Program Grant 350241) and the Aus-tralian Research Council (ARC; Discovery Grant DP0878136).Dr. Whittle is supported by an Australian Research CouncilPostdoctoral Fellowship. Dr. Yucel is supported by an NHMRCClinical Career Development Award (ID: 509345). Ms. Loren-zetti is supported by an Endeavour International Postgradu-ate Research Scholarship (IPRS) and a MelbourneInternational Research Scholarship (MIRS). Prof. Panteliswas supported by a NHMRC Senior Principal Research Fellow-ship (ID: 628386).

The Colonial Foundation, NHMRC, ARC, University of Mel-bourne (providing the MIRS), and Australian Government(providing the IPRS) had no further role in study design; in

the collection, analysis and interpretation of data; in thewriting of the report; and in the decision to submit the paperfor publication.

Conflict of interest

All authors declare that they have no conflicts of interest.

Acknowledgements

Neuroimaging analysis was facilitated by the NeuropsychiatryImaging Laboratory managed by Ms. Bridget Soulsby at theMelbourne Neuropsychiatry Centre and supported by Neu-rosciences Victoria. The authors would like to thank the BrainResearch Institute for support in acquiring the neuroimagingdata and the families who participated in the study.

Appendix A. Supplementary data

Supplementary data associated with this article can befound, in the online version, at doi:10.1016/j.psyneuen.2011.10.004.

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