Disconnectivity between the raphe nucleus and subcortical dopamine-related regions contributes altered salience network in schizophrenia

Schizophrenia Research xxx (xxxx) xxx

Contents lists available at ScienceDirect

Schizophrenia Research journal homepage: www.elsevier.com/locate/schres

Disconnectivity between the raphe nucleus and subcortical dopamine-related regions contributes altered salience network in schizophrenia Shaoqiang Han a, b, Qian Cui d, Xiaonan Guo a, b, Yun-Shuang Fan a, b, Jing Guo a, b, Xiaofen Zong c, Maolin Hu c, Fengmei Lu a, b, Xiaogang Chen c, **, Huafu Chen a, b, * a

The Clinical Hospital of Chengdu Brain Science Institute, MOE Key Lab for Neuroinformation, University of Electronic Science and Technology of China, Chengdu, China b School of Life Science and Technology, Center for Information in Medicine, University of Electronic Science and Technology of China, Chengdu, China c Mental Health Institute of the Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, China d School of Public Administration, University of Electronic Science and Technology of China, Chengdu, China

a r t i c l e i n f o

a b s t r a c t

Article history: Received 22 January 2019 Received in revised form 28 May 2019 Accepted 3 November 2019 Available online xxx

Numerous studies strongly have suggested the significant role of serotonin in the pathomechanism of schizophrenia. However, few studies have directly explored the altered serotonin function in schizophrenia. In the current study, we explored the altered serotonin function in first-episode treatment-naive patients with schizophrenia with resting-state functional magnetic resonance imaging. A total 42 firstepisode treatment-naive patients with schizophrenia and carefully matched healthy controls are included in the study. Considering that the raphe nucleus providing a substantial proportion of the serotonin innervation to the forebrain, the raphe nucleus was chosen as the seed to construct voxelbased functional connectivity (FC) maps. In the results, subcortical dopamine-related regions presented decreased FC with the raphe nucleus, such as the bilateral striatum, pallidum, and thalamus, in patients with schizophrenia. Decreased FC in these regions was significantly correlated with the total negative scores in PANSS. Furthermore, these regions presented with decreased FC connection to salience network. Our results presented that the raphe nucleus played an important role in the dysfunction of subcortical DA-related regions, and contributed to the altered salience network in schizophrenia. Our study emphasized the importance of the raphe nucleus in the pathophysiology of schizophrenia. © 2019 Elsevier B.V. All rights reserved.

Keywords: Schizophrenia Serotonin Raphe nucleus Salience network

1. Introduction As a severe psychiatric disorder affecting nearly 1% of the population, schizophrenia has a severe effect on the individuals’ life and society (Ross et al., 2006; Sawa and Snyder, 2002). However, its precise causes remain unknown. For now, the dopamine (DA) hypothesis receives extensive attention (Carlsson and Lindqvist, 1962; Carter and Pycock, 1980; Saunders et al., 1998), and a mountain of evidence studies have implicated the altered dopaminergic neurotransmission in the occurrence of psychotic symptoms

* Corresponding author. Key Laboratory for NeuroInformation of Ministry of Education, School of Life Science and Technology, University of Electronic Science and Technology of China, Chengdu, 610054, China. ** Corresponding author. E-mail addresses: [email protected] (X. Chen), [email protected] (H. Chen).

(Howes and Murray, 2014). However, dopaminergic dysfunction falls to explain a variety of negative symptoms and cognitive impairments. Typical antipsychotic drugs designed to block striatal D2 deceptors (Creese et al., 1976) confer little effect in improving cognitive function (Davidson et al., 2009; Keefe et al., 1999). Recently, atypical antipsychotic drugs are found to have higher affinity of 5-HT2A than D2 receptors (Stockmeier et al., 1993; Zhang and Bymaster, 1999), and help improve negative symptoms and some cognition functions in schizophrenia (Lowe et al., 2018; Meltzer and Mcgurk, 1999; Nucifora et al., 2017). These phenomena have attracted research attention to the important role of serotonin in the pathophysiology of schizophrenia. Serotonin plays an important role in brain development, memory, mood, and cognitive states (Broman and Fletcher, 1999; Kandel, 2001). Numerous studies have suggested the role of serotonin in the pathogenesis of schizophrenia (Abdolmaleky et al., 2014b; Cheah et al., 2017; Clarke et al., 2004; Meltzer et al., 2003;

https://doi.org/10.1016/j.schres.2019.11.006 0920-9964/© 2019 Elsevier B.V. All rights reserved.

Please cite this article as: Han, S et al., Disconnectivity between the raphe nucleus and subcortical dopamine-related regions contributes altered salience network in schizophrenia, Schizophrenia Research, https://doi.org/10.1016/j.schres.2019.11.006

2

S. Han et al. / Schizophrenia Research xxx (xxxx) xxx

Quednow et al., 2009; Selvaraj et al., 2014). The serotonin function is one of the key biomarkers in diagnosis and therapy of schizophrenia (Abdolmaleky et al., 2014b). Moreover, most atypical antipsychotics have affinity for 5-HT receptors (Quednow et al., 2009). For example, clozapine having high affinity for the 5-HT2C receptor antagonists can directly increase dopamine release in regions like the nucleus accumbens and prefrontal cortex (Di et al., 1998). In recent years, the serotonineglutamate receptor complex in cortical pyramidal neurons have been hypothesized in the altered cortical process in schizophrenia, and represents a promising research direction for the treatment of schizophrenia, especially for negative symptoms and cognitive impairment (Aghajanian and Marek, 1999;
lez-Maeso and Sealfon, 2009; Gonz
alez-Maeso et al., 2008; Gonza Gonz
alezmaeso et al., 2008; Miyamoto et al., 2005; Owen et al., 2016). Even though serotonin function research is an promising opportunity for schizophrenia, only a few studies have directly explored altered serotonin function in schizophrenia (Geyer and Vollenweider, 2008; Selvaraj et al., 2014). Resting-state functional magnetic resonance imaging (fMRI) can be a effective tool to explore the altered serotonin function in schizophrenia and it can provide approaches to detect the functional integrity of specific brain circuits in vivo (Harrison et al., 2009; Martino et al., 2011). In schizophrenia, researchers have achieved great success using fMRI. For example, the striatum, which harbors the largest density of dopamine D2 receptors (Hall et al., 1994; Seeman and Lee, 1975). Its function is heavily modulated by DA (Seeman and Lee, 1975). Using the striatum as seed, studies have found that dysregulation of corticostriatal systems presents a putative risk phenotype for schizophrenia (Fornito et al., 2013), and the dysfunction can be a predictive biomarker for improving the symptoms introduced by antipsychotic treatments (Sarpal et al., 2015). The raphe nucleus providing a substantial proportion of the serotonin innervation to the forebrain (Hornung, 2003; Jacobs and Azmitia, 1992; Pollak et al., 2014). In the same way, the raphe nucleus is the primary seed to explore the altered serotonin function in schizophrenia. Additionally, Beliveau et al. find association between functional connectivity (FC) of raphe nucleus and 5-HTT binding, demonstrating serotonergic contribution to the resting-state signal (Beliveau et al., 2015a). Numerous studies have found that subcortical regions, like the striatum, are under inhibitory control of the serotonergic system. The atypical antipsychotic drugs that have been shown to have higher affinity of 5-HT2A than D2 receptors (Meltzer et al., 1989; Stockmeier et al., 1993; Zhang and Bymaster, 1999) and altered function of the 5HT2A receptors, which are mainly located in the prefrontal cortex (Abdolmaleky et al., 2004, 2014a; Fan and Sklar, 2005; Pehek et al., 2006; Williams et al., 1997). Using the raphe nucleus as seed, the altered serotoninergic function of the posterior cingulated cortex has been found in later-life depression (Ikuta et al., 2017). However, there was no research exploring the altered FC of the raphe nuclei in schizophrenia. In the present study, we explored the aberrant FC of the raphe nucleus in treatment-naive patients with schizophrenia with resting-state FC using a seed-based approach. A total of 42 firstepisode drug-naive patients with schizophrenia and matched 38 controls (such as age, gender and education) were included. Combing evidences mentioned before, we hypothesized the FC between subcortical DA related regions and raphe nuclei would be altered. In addition, FC between prefrontal cortex and raphe nucleus might also be altered.

schizophrenia and 38 matched healthy controls (HCs) were enrolled at the Second Affiliated Hospital of Xinxiang Medical University. Patients included in current study met DSM-IV-TR criteria for schizophrenia (Diagnostic and Statistical Manual of Mental Disorders, fourth edition, text revision, American Psychiatric Association, 2000) and did not have no co-morbid Axis I diagnosis. All patients were independently diagnosed by two welltrained psychiatrists and were interviewed again after six months before a final diagnosis of schizophrenia. To control the BOLD quality, the following quality assurance criteria should also be fulfilled for each participant: i) signal-to-noise ratios greater than 100. The signal-to-noise ratios was obtained by calculating the mean and SD for a given time point across scanning in brain voxels; ii) translational and rotational displacement could not exceed 2.0 mm or 2.0
. At last, 39 patients with schizophrenia and 34 healthy controls were included (Table 1). HCs in our study do not presented any history of psychiatric (or neurological disorders) and medical conditions. In addition, exclusion criterions for healthy controls included: (1) family history of hereditary neurological disorders; (2) loss of consciousness resulted from head injury; (3) alcohol or substance abuse; and (4) any metallic objects in their body. Current study was authorized by the ethics committee of the Second Xiangya Hospital and the Second Affiliated Hospital of Xinxiang Medical University. Written informed consents were gotten from all participants. 2.2. Data acquisition Data were acquired in the Second Affiliated Hospital of Xinxiang Medical University with a Siemens 3T Trio scanner (Siemens Medical Systems, Erlangen, Germany). Clinical assessment was conducted first on the same day before scanning. A total of 240 vol (8 min) of echo-planar images were obtained axially with a echoplanar imaging sequence (EPI) with the following parameters: TR/TE ¼ 2000/30 ms, 33 slices, 64  64 matrix, 90
flip angle, field of view ¼ 220  220 mm2, interslice gap ¼ 0.6 mm, and voxel size ¼ 3.44  3.44  4 mm3. 2.3. Data preprocessing The preprocessing steps were done with Data Processing Assistant for Resting-State fMRI package (DPARSFA, http://www. restfmri.net). The following preprocessing steps were included. Removing the first 10 vol, followed by slice timing and realignment. All images were then resampled into 3  3  3 mm3 and spatial normalized to the standard EPI template, smoothed with a Gaussian kernel of 6 mm full width at half maximum (FWHM). The signal-to-noise ratios was obtained by calculating the mean and SD for a given time point across scanning in brain voxels. Subjects

Table 1 Characteristics of patients with schizophrenia and healthy controls.

Age (years), Mean ± SD Gender, male: female Duration of illness (months), Mean ± SD Alcohol, yes/no Cigarette, yes/no Years of education, Mean ± SD Handness, right/left PANSS positive score PANSS negative score PANSS general score PANSS total score

2. Materials and methods 2.1. Participants

Sch (n ¼ 39)

HC (n ¼ 34)

p

25.00 ± 4.75 24:15 8.13 ± 2.54 6/33 8/31 10.36 ± 2.66 39/0 25.56 ± 3.89 18.54 ± 5.13 48.54 ± 6.47 92.64 ± 11.19

25.12 ± 4.58 21:13 e 7/27 8/26 11.12 ± 2.85 34/0 e e e e

0.91a 0.99b e 0.49 b 0.76b 0.24 a e e e e e

a

Forty-two

first-episode

treatment-naive

patients

with

P- value was obtained by two-tailed two-sample t-test. P- value was obtained by c2 two-tailed test.

b

Please cite this article as: Han, S et al., Disconnectivity between the raphe nucleus and subcortical dopamine-related regions contributes altered salience network in schizophrenia, Schizophrenia Research, https://doi.org/10.1016/j.schres.2019.11.006

S. Han et al. / Schizophrenia Research xxx (xxxx) xxx

would be excluded if signal-to-noise ratios less than 100 or translational and rotational displacement exceeded 2.0 mm or 2.0
. Then images were detrended and band-pass filtered (0.01e0.1 Hz). Nuisance covariates including 24 motion parameters, global mean signal, white matter signal and cerebrospinal fluid signal were regressed out (Satterthwaite et al., 2012). Finally, despking where outliners were detected and replaced with the best estimate using a third-order spline fit (3dDespike) (http://afni.nimh.nih.gov/afni) (Allen et al., 2014). To further control the effect of head motion, we also calculated the mean frame-wise displacement (FD) and compared it between HCs and patients (Guo et al., 2018; Han et al., 2017; Liao et al., 2018). We found no significant difference of mean FD between groups with two sample t-test (p ¼ 0.79). 2.4. FC analyses Raphe nuclei was subdivided into dorsal (DR) and median (MR) raphe nuclei (Beliveau et al., 2015a). The DR and MR were defined as 32 voxels regions (spherical radius of 6 mm) centered at DR (0, 27,-9) and MR (0, 31, 21) (Beliveau et al., 2015a; Han et al., 2019; Ikuta et al., 2017; Kranz et al., 2012). The spatially averaged mean value of each seed was extracted. Voxel-wise FC maps were constructed by calculating the Pearson correlation between mean value of seed signal (DR or MR) and each voxel in the grey matter. Finally, correlation coefficients were transformed to Z scores with Fisher’s Z-transform. 2.5. Statistical analysis Two-tailed two sample t-test was used to determine altered FC of raphe nuclei in patients with schizophrenia compared with HCs. In this step, sex, age, years of education, and mean FD were treated as nuisance covariates. Results reported were corrected with the Gaussian random field (GRF) (voxel-wise p < 0.005 and clusterlevel p < 0.05). 2.6. Relation with clinical symptoms To investigate the association between altered connections and clinical symptom severity, we extracted the altered FC of peak coordinates with a spherical radius of 6 mm. The spatial mean values of altered FC was then correlated with PANSS scores with Pearson correlation. 2.7. Validation analysis Previous studies have found that global signal regression affects the FC maps of fMRI (Murphy et al., 2009), especially for schizophrenia (Yang et al., 2014). Therefore, we recalculated our results with no global signal regression. 3. Results 3.1. Clinical effects As shown in Table 1, there was no significant difference between HCs and patients in terms of sociodemographic characteristics such as age, gender and education. 3.2. FC analysis 3.2.1. Decreased functional connectivity of DR and MR in schizophrenia We compared the FC maps of the DR and MR between HCs and patients with schizophrenia. As the results, FC of the DR and MR was decreased in patients. Specifically, bilateral striatum presented

3

decreased FC in terms of connection to the DR. The right thalamus, right caudate, and right putamen presented decreased FC connected to the MR (Fig. 1, Table 1 in supplement results). Considering aberrance was limited to subcortical DA-related regions, we further explored the whole altered FC pathway of the raphe nucleus. The peak coordinates of each cluster that consistently presented significant difference with and without global signal regression (see validation analysis) were chosen. Afterward, the right putamen (27, 6, 3), left pallidum (21, 6, 6), and right thalamus (18, 15, 3) were chosen. The whole brain voxel-based FC maps were constructed using these regions as seeds, and then compared between patients and HCs using two-sample T test. In the results, salience network (SN) presented decreased FC with salience networks, including the bilateral anterior insular cortex, left anterior cingulate cortex, left middle prefrontal cortex, and subcortical DA-related regions (voxel-wise p < 0.005, clusterlevel p < 0.05, GRF corrected) (Fig. 2, Table 2 in supplement results). 3.2.2. Decreased functional connectivity of the raphe nucleus was correlated with the total negative scores To explore the relationship between the dysfunction of raphe nucleus and patients’ symptoms, we examined the correlation between the altered FC strength and total positive and negative PANSS scores. In the results, the strength of the connection linking the right putamen and the DR (r ¼ 0.513, p < 0.001 uncorrected) and the connection linking the left pallidum was significantly correlated with total negative scores (r ¼ 0.363, p ¼ 0.023 uncorrected) (Fig. 3). 3.2.3. Validation results Most of differences found in our study could be replicated without global signal regression (supplemental results). The decreased FC between the right putamen and the DR remained significantly correlated with the total negative score in PANSS without global signal regression (r ¼ 0.416, p ¼ 0.009 uncorrected). Moreover, the decreased FC between the left pallidum and the DR was nearly significantly correlated to the total negative score in PANSS without global signal regression (r ¼ 0.290, p ¼ 0.073 uncorrected). 4. Discussion This current study is the first time that altered FC is examined in the raphe nucleus of first-episode treatment-naive patients with resting-state seed-based FC. In the results, subcortical DA-related regions presented decreased FC with the raphe nucleus. Specially, the bilateral striatum presented with decreased FC connection to the DR. The right thalamus, right caudate, and right putamen presented with decreased FC connection to the MR. Moreover, the decreased FC of these regions was significantly correlated with the total negative scores in PANSS. In addition, the striatum, pallidum, and thalamus further presented with decreased FC connection to SN networks. These results demonstrated that the raphe nucleus is important in the dysfunction of subcortical DA-related regions and contributes to the aberrant salience in schizophrenia. For the first time, the dysfunctional connectivity of the raphe nucleus is presented to have an importance in the pathophysiology of schizophrenia. Our research found that the FCs between the raphe nucleus and subcortical DA-related regions were decreased in patients with schizophrenia. The striatum receives an input from serotonergic neurons originating from the raphe nuclei (Azmitia and Segal, 1978). Systems originating from the ventral tegmental area (VTA) are under phasic and tonic inhibitory control by the serotonergic system (Di et al., 2002; Gudelsky et al., 1994; Vincenzo et al., 2002). A large body of evidence demonstrate the decreased inhibition of

Please cite this article as: Han, S et al., Disconnectivity between the raphe nucleus and subcortical dopamine-related regions contributes altered salience network in schizophrenia, Schizophrenia Research, https://doi.org/10.1016/j.schres.2019.11.006

4

S. Han et al. / Schizophrenia Research xxx (xxxx) xxx

Fig. 1. Significant decreased functional connectivity of raphe nuclei in patients, comparing with HC. The left letters indicate the seeds of functional connectivity and blue regions in the right brain slices present the location of difference (GRF corrected). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 2. Significant decreased functional connectivity of subcortical DA related regions in patients, comparing with HC. The left letters indicate the seeds of functional connectivity and blue regions in the right brain slices present the location of difference (GRF corrected). Note: R.Put, right putamen; L.Pal, left pallidum; R.Tha, right thalamus. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

dopamine release and its pivotal role in schizophrenia (Castensson et al., 2003, 2010; Dracheva et al., 2010; Lewis et al., 1999; Olaya et al., 2018; Yao et al., 2018). Combining the our findings and the serotonergic contribution to the resting-state signal (Beliveau et al., 2015a), the decreased FC between the raphe nuclei and subcortical DA-related regions hinted on the decreased inhibitory control on the DA system. However, the hypothesis needs robust evidence. Another finding was that the decreased FC between subcortical DA-related regions and the raphe nucleus was associated with negative symptoms. This association might reflect deficit in anticipatory motivation of patients with schizophrenia. Deficit in anticipatory motivation has been linked to striatal dysfunction

resulting from increased striatal D2 receptors (D2R) density or occupancy (Sorg et al., 2013a). Furthermore, the mouse in which D2Rs were selectively overexpressed in the striatum displayed deficient motivation (Simpson and Kellendonk, 2016). An acute increase in striatal D2Rs might be respond to the deficit in anticipatory motivation in patients with schizophrenia. These results might be related to deficiency in anticipatory motivation in patients with schizophrenia. Relative to HCs, treatment-naive patients demonstrated decreased functional connectivity between the striatum and SN. This might have resulted from elevated dopamine levels that worsened the signal-to-noise ratio of spontaneous brain activity in

Please cite this article as: Han, S et al., Disconnectivity between the raphe nucleus and subcortical dopamine-related regions contributes altered salience network in schizophrenia, Schizophrenia Research, https://doi.org/10.1016/j.schres.2019.11.006

S. Han et al. / Schizophrenia Research xxx (xxxx) xxx

5

Fig. 3. The negative symptoms was correlated with decreased FC of the raphe nucleus. The y-coordinate meant the total negative score. The x-coordinate meant z-score of FC in patients.

the striatum (Sorg et al., 2013b). In schizophrenia, frontostriatal network level alterations have been suggested to be a putative risk endophenotype for schizophrenia (Dandash et al., 2014; Fornito et al., 2013; Hu et al., 2016). The anterior insula cortex (AIC) and dorsal anterior cingulate cortex (dACC) have been thought to have roles in salience processing (Uddin, 2015). These decreased connections found in our study might embody aberrant salience attribution, and these are consistent with the theory of ‘dysconnectivity’ hypotheses in schizophrenia (Friston and Frith, 1995; Zalesky et al., 2010, 2015). We did not observe significant difference of FC between prefrontal regions and the raphe nucleus. Several reasons exist: altered function of the 5-HT2A receptor has been associated with schizophrenia (Abdolmaleky et al., 2004, 2014a; Fan and Sklar, 2005; Pehek et al., 2006; Williams et al., 1997), but this remains contro
pezfigueroa et al., 2004). Another reason might be that versial (Lo other studies used different ligands (Selvaraj et al., 2014). The conclusion that a significant association exists among DR, MR FC, and regional 5-HTT binding in a previous study (Beliveau et al., 2015a) might be limited by the ligand used. Another reason might be that effects of chronic antipsychotic medication may contribute to these different results (Bantick et al., 2001). The last reason might be the relatively small sample size, and future research could use a larger sample to validate the results.

These results demonstrate that the raphe nucleus is important in the dysfunction of subcortical DA-related regions, and contribute to the aberrant salience in schizophrenia. Our study emphasizes the importance of the raphe nucleus in the pathophysiology of schizophrenia.

4.1. Limitation

Abdolmaleky, H.M., Faraone, S.V., Glatt, S.J., Tsuang, M.T., 2004. Meta-analysis of association between the T102C polymorphism of the 5HT2a receptor gene and schizophrenia. Schizophr. Res. 67 (1), 53. Abdolmaleky, H.M., Nohesara, S., Ghadirivasfi, M., Lambert, A.W., Ahmadkhaniha, H., Ozturk, S., Chen, K.W., Shafa, R., Mostafavi, A., Thiagalingam, S., 2014. DNA hypermethylation of serotonin transporter gene promoter in drug naive patients with schizophrenia. Schizophr. Res. 152 (2e3), 373. Abdolmaleky, H.M., Nohesara, S., Ghadirivasfi, M., Lambert, A.W., Ahmadkhaniha, H., Ozturk, S., Wong, C.K., Shafa, R., Mostafavi, A., Thiagalingam, S., 2014. DNA hypermethylation of serotonin transporter gene promoter in drug naive patients with schizophrenia. Schizophr. Res. 152 (2e3), 373e380. Aghajanian, G.K., Marek, G.J., 1999. Serotonin|[ndash]|Glutamate interactions: a new target for antipsychotic drugs. Neuropsychopharmacology 21 (6), S122eS133. Allen, E.A., Damaraju, E., Plis, S.M., Erhardt, E.B., Eichele, T., Calhoun, V.D., 2014. Tracking whole-brain connectivity dynamics in the resting state. Cerebr. Cortex 24 (3), 663. Azmitia, E.C., Segal, M., 1978. Azmitia EC, Segal M. Autoradiographic analysis of differential ascending projections of dorsal and median raphe nuclei in rat. J Comp Neurol 179: 641-667. J. Comp. Neurol. 179 (3), 641e667. Bantick, R.A., Deakin, J.F., Grasby, P.M., 2001. The 5-HT1A receptor in schizophrenia: a promising target for novel atypical neuroleptics? J. Psychopharmacol. 15 (1), 37. Beliveau, V., Svarer, C., Frokjaer, V.G., Knudsen, G.M., Greve, D.N., Fisher, P.M., 2015. Functional connectivity of the dorsal and median raphe nuclei at rest. Neuroimage 116, 187e195. Beliveau, V., Svarer, C., Frokjaer, V.G., Knudsen, G.M., Greve, D.N., Fisher, P.M., 2015. Functional connectivity of the dorsal and median raphe nuclei at rest.

One main limitation in the present study was the modest sample size. A larger sample size would better elaborate on the FC between the raphe nucleus and prefrontal regions in patients with schizophrenia. Another limitation was that our results did not have replications in another cohorts of patients with schizophrenia. Future studies are suggested to use larger sample sizes to validate our results. The last, there are a lot non-serotonergic neurons in DR and MR, future studies could directly manipulate 5-HT (such as tryptophan deletion or 5-HTT inhibition) to explore aspects of the serotonergic contribution to raphe FC (Beliveau et al., 2015b). 5. Conclusion The current study shows for the first time the altered FC of the DR and MR in patients with schizophrenia with resting-state FC using a seed-based approach. In the results, subcortical DA-related regions presented with decreased FC with the raphe nucleus. Decreased FC of these regions was significantly correlated with the total negative scores in PANSS. The striatum, pallidum, and thalamus presented with decreased FC connection to SN networks.

Declaration of competing interests All authors declared no conflicts of interest. Acknowledgments This work was supported by grants from the National Natural Science Foundation of China (61533006, U1808204, 81271484,81771919 and 81471361), Sichuan Science and Technology Program (2018TJPT0016), China Postdoctoral Science Foundation Grant (Grant No. 2019M653383). Appendix A. Supplementary data Supplementary data to this article can be found online at https://doi.org/10.1016/j.schres.2019.11.006. References

Please cite this article as: Han, S et al., Disconnectivity between the raphe nucleus and subcortical dopamine-related regions contributes altered salience network in schizophrenia, Schizophrenia Research, https://doi.org/10.1016/j.schres.2019.11.006

6

S. Han et al. / Schizophrenia Research xxx (xxxx) xxx

Neuroimage 116 (2), 187e195. Broman, S.H., Fletcher, J.M., 1999. The changing nervous system: neurobehavioral consequences of early brain disorders. Contemp. Psychol. Rev. Books. 46 (4), 353e355. Carlsson, A., Lindqvist, M., 1962. In-vivo decarboxylation of alpha-methyl DOPA and alpha-methyl metatyrosine. Acta Physiol. Scand. 54 (1), 87e94. Carter, C.J., Pycock, C.J., 1980. Behavioural and biochemical effects of dopamine and noradrenaline depletion within the medial prefrontal cortex of the rat. Brain Res. 192 (192), 163e176. Castensson, A., Aring, Berg, K., Mccarthy, S., Saetre, P., Andersson, B., Jazin, E., 2010. Serotonin receptor 2C (HTR2C) and schizophrenia: examination of possible medication and genetic influences on expression levels. Am J Med Genet B Neuropsychiatr Genet 134B (1), 84e89. Castensson, A., Emilsson, L., Sundberg, R., Jazin, E., 2003. Decrease of serotonin receptor 2C in schizophrenia brains identified by high-resolution mRNA expression analysis. Biol. Psychiatry 54 (11), 1212e1221. Cheah, S.Y., Lawford, B.R., Young, R.M., Morris, C.P., Voisey, J., 2017. mRNA expression and DNA methylation analysis of serotonin receptor 2A (HTR2A) in the human schizophrenic brain. Genes 8 (1), 11. Clarke, H.F., Dalley, J.W., Crofts, H.S., Robbins, T.W., Roberts, A.C., 2004. Cognitive inflexibility after prefrontal serotonin depletion. Science 304 (5672), 878e880. Creese, I., Burt, D.R., Snyder, S.H., 1976. Dopamine receptor binding predicts clinical and pharmacological potencies of antischizophrenic drugs. J. Neuropsychiatry Clin. Neurosci. 192 (4238), 481e483. Dandash, O., Fornito, A., Lee, J., Keefe, R.S.E., Chee, M.W.L., Adcock, R.A., Pantelis, C., Wood, S.J., Harrison, B.J., 2014. Altered striatal functional connectivity in subjects with an at-risk mental state for psychosis. Schizophr. Bull. 40 (4), 904e913. Davidson, M., Galderisi, S., Weiser, M., Werbeloff, N., Fleischhacker, W.W., Keefe, R.S., Boter, H., Keet, I.P., Prelipceanu, D., Rybakowski, J.K., 2009. Cognitive effects of antipsychotic drugs in first-episode schizophrenia and schizophreniform disorder: a randomized, open-label clinical trial (EUFEST). Am. J. Psychiatry 166 (6), 675. Di, M.V., Cacchio, M., Di, G.C., Esposito, E., 2002. Role of serotonin(2C) receptors in the control of brain dopaminergic function. Pharmacol., Biochem. Behav. 71 (4), 727e734. Di, M.V., Di, G.G., Di, M.M., Esposito, E., 1998. Selective blockade of serotonin2C/2B receptors enhances dopamine release in the rat nucleus accumbens. Neuropharmacology 37 (2), 265. Dracheva, S., Elhakem, S.L., Marcus, S.M., Siever, L.J., Mcgurk, S.R., Haroutunian, V., 2010. RNA editing and alternative splicing of human serotonin 2C receptor in schizophrenia. J. Neurochem. 87 (6), 1402e1412. Fan, J.B., Sklar, P., 2005. Meta-analysis reveals association between serotonin transporter gene STin2 VNTR polymorphism and schizophrenia. Mol. Psychiatry 10 (10), 928. Fornito, A., Harrison, B.J., Goodby, E., Dean, A., Ooi, C., Nathan, P.J., Lennox, B.R., Jones, P.B., Suckling, J., Bullmore, E.T., 2013. Functional dysconnectivity of corticostriatal circuitry as a risk phenotype for psychosis. Jama Psychiatry 70 (11), 1143e1151. Friston, K.J., Frith, C.D., 1995. Schizophrenia: a disconnection syndrome? Clin. Neurosci. 3 (2), 89e97. Geyer, M.A., Vollenweider, F.X., 2008. Serotonin research: contributions to understanding psychoses. Trends Pharmacol. Sci. 29 (9), 445e453.
lez-Maeso, J., Ang, R., Yuen, T., Chan, P., Weisstaub, N.V., Lo
pez-Gime
nez, J.F., Gonza Zhou, M., Okawa, Y., Callado, L.F., Milligan, G., 2008. Identification of a serotonin/glutamate receptor complex implicated in psychosis. Nature 452 (7183), 93.
lez-Maeso, J., Sealfon, S.C., 2009. Psychedelics and schizophrenia. Trends Gonza Neurosci. 32 (4), 225e232.
lezmaeso, J., Ang, R.L., Yuen, T., Chan, P., Weisstaub, N.V., Lo
pezgime
nez, J.F., Gonza Zhou, M., Okawa, Y., Callado, L.F., Milligan, G., 2008. Identification of a serotonin/glutamate receptor complex implicated in psychosis. Nature 452 (7183), 93. Gudelsky, G.A., Yamamoto, B.K., Nash, J.F., 1994. Potentiation of 3,4methylenedioxymethamphetamine-induced dopamine release and serotonin neurotoxicity by 5-HT2 receptor agonists. Eur. J. Pharmacol. 264 (3), 325. Guo, X., Duan, X., Suckling, J., Chen, H., Liao, W., Cui, Q., Chen, H., 2018. Partially Impaired Functional Connectivity States between Right Anterior Insula and Default Mode Network in Autism Spectrum Disorder. Human Brain Mapping. Hall, H., Sedvall, G., Magnusson, O., Kopp, J., Halldin, C., Farde, L., 1994. Distribution of D1- and D2-dopamine receptors, and dopamine and its metabolites in the human brain. Neuropsychopharmacology 11 (4), 245. Han, S., He, Z., Duan, X., Tang, Q., Chen, Y., Yang, Y., Pang, Y., Nan, X., Cui, Q., Chen, H., 2019. Dysfunctional connectivity between raphe nucleus and subcortical regions presented opposite differences in bipolar disorder and major depressive disorder. Prog. Neuro Psychopharmacol. Biol. Psychiatry 92, 76e82. Han, S., Zong, X., Hu, M., Yu, Y., Wang, X., Long, Z., Wang, Y., Chen, X., Chen, H., 2017. Frequency-selective alteration in the resting-state corticostriatal-thalamocortical circuit correlates with symptoms severity in first-episode drug-naive patients with schizophrenia. Schizophr. Res. 189, 175e180.
pezsol
ndezribas, R., Harrison, B.J., Sorianomas, C., Pujol, J., Ortiz, H., Lo a, M., Herna Deus, J., Alonso, P., Yücel, M., Pantelis, C., 2009. Altered corticostriatal functional connectivity in obsessive-compulsive disorder. Neuroimage 47 (11). S49-S49. Hornung, J.P., 2003. The human raphe nuclei and the serotonergic system. J. Chem. Neuroanat. 26 (4), 331e343. Howes, O.D., Murray, R.M., 2014. Schizophrenia: an integrated sociodevelopmentalcognitive model. Lancet 383 (9929), 1677e1687. Hu, M., Zong, X., Zheng, J., Mann, J.J., Li, Z., Pantazatos, S.P., Li, Y., Liao, Y., He, Y.,

Zhou, J., 2016. Risperidone-induced topological alterations of anatomical brain network in first-episode drug-naive schizophrenia patients: a longitudinal diffusion tensor imaging study. Psychol. Med. 46 (12), 1. Ikuta, T., Matsuo, K., Harada, K., Nakashima, M., Hobara, T., Higuchi, N., Higuchi, F., Otsuki, K., Shibata, T., Watanuki, T., 2017. Disconnectivity between dorsal raphe nucleus and posterior cingulate cortex in later life depression. Front. Aging Neurosci. 9, 236. Jacobs, B.L., Azmitia, E.C., 1992. Structure and function of the brain serotonin system. Physiol. Rev. 72 (1), 165e229. Kandel, E.R., 2001. The molecular biology of memory storage: a dialogue between genes and synapses. Science 21 (5), 565e611. Keefe, R.S., Silva, S.G., Perkins, D.O., Lieberman, J.A., 1999. The effects of atypical antipsychotic drugs on neurocognitive impairment in schizophrenia: a review and meta-analysis. Schizophr. Bull. 25 (2), 201. Kranz, G.S., Hahn, A., Savli, M., Lanzenberger, R., 2012. Challenges in the differentiation of midbrain raphe nuclei in neuroimaging research. Proc. Natl. Acad. Sci. U.S.A. 109 (29), E2000.
pezfigueroa, A.L., Norton, C.S., Lo
pezfigueroa, M.O., Armellinidodel, D., Burke, S., Lo
pez, J.F., Watson, S.J., 2004. Serotonin 5-HT1A, 5-HT1B, and 5-HT2A Akil, H., Lo receptor mRNA expression in subjects with major depression, bipolar disorder, and schizophrenia. Biol. Psychiatry 55 (3), 225e233. Lewis, R., Kapur, S., Jones, C., Dasilva, J., Brown, G.M., Wilson, A.A., Houle, S., Zipursky, R.B., 1999. Serotonin 5-HT2 receptors in schizophrenia: a PET study using [18F]setoperone in neuroleptic-naive patients and normal subjects. Am. J. Psychiatry 156 (1), 72e78. Liao, W., Fan, Y.-S., Yang, S., Li, J., Duan, X., Cui, Q., Chen, H., 2018. Preservation effect: cigarette smoking acts on the dynamic of influences among unifying neuropsychiatric triple networks in schizophrenia. Schizophr. Bull. 45 (6), 1242-1240. Lowe, P., Krivoy, A., Porffy, L., Henriksdottir, E., Eromona, W., Shergill, S.S., 2018. When the drugs don’t work: treatment-resistant schizophrenia, serotonin and serendipity. Ther. Adv. Psychopharmacol. 8 (1), 63e70. Martino, A.D., Kelly, C., Grzadzinski, R., Zuo, X.N., Mennes, M., Mairena, M.A., Lord, C., Castellanos, F.X., Milham, M.P., 2011. Aberrant striatal functional connectivity in children with autism. Biol. Psychiatry 69 (9), 847e856. Meltzer, H.Y., Li, Z., Kaneda, Y., Ichikawa, J., 2003. Serotonin receptors: their key role in drugs to treat schizophrenia. Prog. Neuropsychopharmacol. Biol. Psychiatry 27 (7), 1159e1172. Meltzer, H.Y., Matsubara, S., Lee, J.C., 1989. Classification of typical and atypical antipsychotic drugs on the basis of dopamine D-1, D-2 and serotonin2 pKi values. J. Pharmacol. Exp. Ther. 251 (1), 238e246. Meltzer, H.Y., Mcgurk, S.R., 1999. The effects of clozapine, risperidone, and olanzapine on cognitive function in schizophrenia. Schizophr. Bull. 25 (2), 233e255. Miyamoto, S., Duncan, G.E., Marx, C.E., Lieberman, J.A., 2005. Treatments for schizophrenia: a critical review of pharmacology and mechanisms of action of antipsychotic drugs. Mol. Psychiatry 10 (1), 79. Murphy, K., Birn, R.M., Handwerker, D.A., Jones, T.B., Bandettini, P.A., 2009. The impact of global signal regression on resting state correlations: are anticorrelated networks introduced? Neuroimage 44 (3), 893e905. Nucifora, F.C., Mihaljevic, M., Lee, B.J., Sawa, A., 2017. Clozapine as a model for antipsychotic development. Neurotherapeutics 14 (3), 750e761. Olaya, J.C., Heusner, C.L., Matsumoto, M., Sinclair, D., Kondo, M.A., Karl, T., Shannon, W.C., 2018. Overexpression of neuregulin 1 type III confers Hippocampal mRNA alterations and schizophrenia-like behaviors in mice. Schizophr. Bull. 44 (4), 865e875. Owen, M.J., Sawa, A., Mortensen, P.B., 2016. Schizophrenia. Lancet 388 (10039), 86e97. Pehek, E.A., Nocjar, C., Roth, B.L., Byrd, T.A., Mabrouk, O.S., 2006. Evidence for the preferential involvement of 5-HT2A serotonin receptors in stress- and druginduced dopamine release in the rat medial prefrontal cortex. Neuropsychopharmacology 31 (2), 265e277. Pollak, nbsp, Dorocic, Iskra, Fürth, Daniel, Xuan, Yang, Johansson, Yvonne, Pozzi, 2014. A whole-brain atlas of inputs to serotonergic neurons of the dorsal and median raphe nuclei. Neuron 83 (3), 663e678. Quednow, B.B., Geyer, M.A., Halberstadt, A.L., 2009. Serotonin and Schizophrenia. Ross, C.A., Margolis, R.L., Reading, S.A.J., Pletnikov, M., Coyle, J.T., 2006. Neurobiology of schizophrenia. Neuron 52 (1), 139e153. Sarpal, D.K., Robinson, D.G., Lencz, T., Argyelan, M., Ikuta, T., Karlsgodt, K., Gallego, J.A., Kane, J.M., Szeszko, P.R., Malhotra, A.K., 2015. Antipsychotic treatment and functional connectivity of the striatum in first-episode schizophrenia. Jama Psychiatry 72 (1), 5. Satterthwaite, T.D., Wolf, D.H., Loughead, J., Ruparel, K., Elliott, M.A., Hakonarson, H., Gur, R.C., Gur, R.E., 2012. Impact of in-scanner head motion on multiple measures of functional connectivity: relevance for studies of neurodevelopment in youth. Neuroimage 60 (1), 623e632. Saunders, R.C., Kolachana, B.S., Bachevalier, J., Weinberger, D.R., 1998. Neonatal lesions of the medial temporal lobe disrupt prefrontal cortical regulation of striatal dopamine. Nature 393 (6681), 169e171. Sawa, A., Snyder, S.H., 2002. Schizophrenia: diverse approaches to a complex disease. Science 296 (5568), 692e695. Seeman, P., Lee, T., 1975. Antipsychotic drugs: direct correlation between clinical potency and presynaptic action on dopamine neurons. Science 188 (4194), 1217. Selvaraj, S., Arnone, D., Cappai, A., Howes, O., 2014. Alterations in the serotonin system in schizophrenia: a systematic review and meta-analysis of postmortem and molecular imaging studies. Neurosci. Biobehav. Rev. 45, 233e245. Simpson, E.H., Kellendonk, C., 2016. Insights about striatal circuit function and schizophrenia from a mouse model of D2 receptor upregulation *. Biol.

Please cite this article as: Han, S et al., Disconnectivity between the raphe nucleus and subcortical dopamine-related regions contributes altered salience network in schizophrenia, Schizophrenia Research, https://doi.org/10.1016/j.schres.2019.11.006

S. Han et al. / Schizophrenia Research xxx (xxxx) xxx Psychiatry 81 (1). Sorg, C., Manoliu, A., Neufang, S., Myers, N., Peters, H., Schwerth~ a¶Ffer, D., Scherr, M., ~¼Hlau, M., Zimmer, C., Drzezga, A., 2013. Increased intrinsic brain activity in Ma the striatum reflects symptom dimensions in schizophrenia. Schizophr. Bull. 39 (2), 387e395. €ffer, D., Scherr, M., Sorg, C., Manoliu, A., Neufang, S., Myers, N., Peters, H., Schwertho Mühlau, M., Zimmer, C., Drzezga, A., 2013. Increased intrinsic brain activity in the striatum reflects symptom dimensions in schizophrenia. Schizophr. Bull. 39 (2), 387e395. Stockmeier, C.A., Dicarlo, J.J., Zhang, Y., Thompson, P., Meltzer, H.Y., 1993. Characterization of typical and atypical antipsychotic drugs based on in vivo occupancy of serotonin2 and dopamine2 receptors. J. Pharmacol. Exp. Ther. 266 (3), 1374e1384. Uddin, L.Q., 2015. Salience processing and insular cortical function and dysfunction. Nat. Rev. Neurosci. 16 (1), 55e61. Vincenzo, D.M., Marisa, C., Camillo, D.G., Ennio, E., 2002. Role of serotonin(2C) receptors in the control of brain dopaminergic function. Pharmacol. Biochem. Behav. 71 (4), 727e734. €then, M., Owen, M.J., 1997. Meta-analysis of association Williams, J., Mcguffin, P., No

7

between the 5-HT2a receptor T102C polymorphism and schizophrenia. EMASS collaborative group. European multicentre association study of schizophrenia. Lancet 349 (9060), 1221. Yang, G.J., Murray, J.D., Repovs, G., Cole, M.W., Savic, A., Glasser, M.F., Pittenger, C., Krystal, J.H., Wang, X.J., Pearlson, G.D., 2014. Altered global brain signal in schizophrenia. Proc. Natl. Acad. Sci. U.S.A. 111 (20), 7438e7443. Yao, P., Mccorvy, J.D., Harpsøe, K., Lansu, K., Yuan, S., Popov, P., Lu, Q., Pu, M., Tao, C., Nikolajsen, L.F., 2018. 5-HT2C receptor structures reveal the structural basis of GPCR polypharmacology. Cell 172 (4). Zalesky, A., Fornito, A., Seal, M.L., Cocchi, L., Westin, C.F., Bullmore, E.T., Egan, G.F., Pantelis, C., 2010. Disrupted axonal fiber connectivity in schizophrenia. Biol. Psychiatry 69 (1), 80e89. Zalesky, A., Pantelis, C., Cropley, V., Fornito, A., Cocchi, L., Mcadams, H., Clasen, L., Greenstein, D., Rapoport, J.L., Gogtay, N., 2015. Delayed development of brain connectivity in adolescents with schizophrenia and their unaffected siblings. JAMA . Psychiatr. 72 (9), 173e174. Zhang, W., Bymaster, F.P., 1999. The in vivo effects of olanzapine and other antipsychotic agents on receptor occupancy and antagonism of dopamine D1, D2, D3, 5HT2A and muscarinic receptors. Psychopharmacology 141 (3), 267.

Please cite this article as: Han, S et al., Disconnectivity between the raphe nucleus and subcortical dopamine-related regions contributes altered salience network in schizophrenia, Schizophrenia Research, https://doi.org/10.1016/j.schres.2019.11.006