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Rostral anterior cingulate glutamate predicts response to subcallosal deep brain stimulation for resistant depression
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Rostral anterior cingulate glutamate predicts response to subcallosal deep brain stimulation for resistant depression Darren L. Clark , Frank P. MacMaster , Elliot C. Brown , Zelma H.T. Kiss , Rajamannar Ramasubbu PII: DOI: Reference:
S0165-0327(19)33129-5 https://doi.org/10.1016/j.jad.2020.01.058 JAD 11507
To appear in:
Journal of Affective Disorders
Received date: Revised date: Accepted date:
8 November 2019 9 January 2020 14 January 2020
Please cite this article as: Darren L. Clark , Frank P. MacMaster , Elliot C. Brown , Zelma H.T. Kiss , Rajamannar Ramasubbu , Rostral anterior cingulate glutamate predicts response to subcallosal deep brain stimulation for resistant depression, Journal of Affective Disorders (2020), doi: https://doi.org/10.1016/j.jad.2020.01.058
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Highlights Sham-controlled SCC-DBS trial showed negative results in patients with TRD Biomarker-stratified patient selection may improve the SCC-DBS outcomes Lower baseline rACC-Glu (1H -MRS) predicted response at 6 months in 16 TRD patients Excluding comorbid bipolar disorder patients improved prediction Pre-DBS rACC-Glu level may have potentials as a biomarker for patient selection
Rostral anterior cingulate glutamate predicts response to subcallosal deep brain stimulation for resistant depression
Darren L. Clark1-4, Frank P. MacMaster1,3-6, Elliot C. Brown1-4, Zelma H. T. Kiss2-4, Rajamannar Ramasubbu1,3,4. 1
Department of Psychiatry, University of Calgary, Calgary, AB, Canada
2 3
Department of Clinical Neuroscience, University of Calgary, AB, Canada
Mathison Centre for Mental Health Research and Education, University of Calgary,
Calgary, AB, Canada 4
Hotchkiss Brain Institute, University of Calgary, Calgary, AB, Canada
5
Department of Radiology, University of Calgary, Calgary, AB, Canada
6
Child and Adolescent Imaging Research Program, Alberta Children’s Hospital, Calgary,
AB, Canada
*CORRESPONDENCE: Rajamannar Ramasubbu MD, Department of Psychiatry/ Clinical Neurosciences, University of Calgary, Mathison Centre for Mental Health Research and Education, TRW building, Room 4D64, 3280 Hospital Drive NW, Calgary, Alberta, T2N4Z6 Tel +1 403 210 6890; Fax +1 403 210 9114, Email: [email protected].
Keywords: deep brain stimulation, treatment-resistant depression, glutamate, glutamine, anterior cingulate cortex, magnetic resonance imaging
Abstract Background: Deep brain stimulation (DBS) of the subcallosal cingulate (SCC) provided benefit for treatment-resistant depression (TRD) in open-label studies but failed in a
recent randomized sham-controlled trial. Informed patient selection, based on reliable biomarkers, is needed to optimize outcome. We investigated if rostral anterior cingulate (rACC) glutamate/glutamine concentration could serve as a potential biomarker of response. Methods: Sixteen adults with TRD (Major Depression; MDD = 14; Bipolar Depression; BD =2) underwent proton magnetic resonance spectroscopy using a short-echo proton spectroscopy with a voxel placed in the rACC, prior to DBS. Improvement in depression was assessed using the 17-item Hamilton Rating Scale for Depression (HRSD). Glutamate and glutamine concentrations at baseline in the rACC were examined in relation to clinical outcomes at six months. Results: Lower baseline glutamate predicted significant reduction in HRSD scores in all patients (p=0.018), and predicted both HDRS reduction (p=0.002) and 6-month response outcome in MDD patients (p=0.013). Neither baseline glutamine, nor glutamine/glutamate ratio significantly related to outcome nor symptom improvement. Limitations: Our study was limited by sample size, though it is large for a DBS study. We measured from a single voxel in the brain, so we cannot be certain our findings are specific to the rACC. Conclusions: These preliminary results suggest that baseline rACC-glutamate concentration could serve as a response-predictive biomarker for SCC-DBS, particularly in patients with resistant major depression. If our findings are replicated and validated, rACC-glutamate may provide a basis to prospectively select TRD patients to improve likelihood of response to SCC-DBS.
1. Introduction Subcallosal cingulate (SCC) deep brain stimulation (DBS) is a promising investigational intervention for treatment-resistant depression (TRD) with 6-month response rates around 50% in open-label trials (Dandekar et al., 2018). Unfortunately, a double-blind randomized, controlled trial of SCC-DBS failed to demonstrate difference between sham and active DBS at 6-months (Holtzheimer et al., 2017). This failure has been
attributed to various factors, including inconsistent patient selection. DBS is invasive, costly and associated with significant risk. Informed patient selection strategies, based on reliable response biomarkers, would defray costs and limit the risks of invasive surgery to those most likely to benefit. The rostral anterior cingulate (rACC; Brodmann area (BA)24) lies anterior and dorsal to the SCC target region and has emerged as a promising biomarker of illness and predictor of treatment response in depression (reviewed in Pizzagalli, 2011 (Pizzagalli, 2011)). Altered glutamatergic signaling in the rACC is implicated in MDD pathology. Reduced glutamate, glutamine or combined glutamate-glutamine (Glx) levels, measured with proton magnetic resonance spectroscopy (MRS), have been described in the rACC of adults (Auer et al., 2000; Merkl et al., 2011) and youths (Mirza et al., 2004) with MDD. Lower rACC glutamate levels have been related to anhedonia, both directly through relationship with subjective emotional rating and indirectly via correlation with reduced negative bold response to emotional stimuli (Walter et al., 2009). It was also related to reduced resting functional connectivity of the rACC to the anterior insula, a network required for proper saliency processing and switching between default-mode network and executive function (Horn et al., 2010). Altered glutamate levels in the rACC might represent a failure of the greater ACC circuit to properly regulate emotional responses, and facilitate task-based network switching, contributing to emotional dysregulation and perseverative rumination. Furthermore, The rACC is tightly interconnected to the SCC (Heilbronner and Haber, 2014; Palomero-Gallagher et al., 2008) and the function of these regions is highly (though often inversely) correlated (Chiba et al., 2001; Johansen-Berg et al., 2008; Pizzagalli, 2011). Stimulation of the local SCC with DBS would likely alter the broader ACC function, either directly via cingulate white matter activation (Riva-Posse et al., 2018; 2014) or indirectly via local SCC activity changes impacting the function of rACC via reciprocal connections (Chiba et al., 2001; Heilbronner and Haber, 2014; Johansen-Berg et al., 2008). Although direct evidence Is lacking, preclinical studies suggest that DBS may modulate excitatory glutamatergic neurotransmission (Ross et al., 2016). In addition, pre-treatment glutamatergic levels in
the rACC predicted response and/or were normalized after electroconvulsive therapy (ECT) in patients with TRD (Pfleiderer et al., 2003; Zhang et al., 2013), providing further support for the potential utility of rACC glutamatergic signaling as a predictor and response biomarker for SCC-DBS in TRD patients. We hypothesized that lower rACC glutamate or glutamine levels would predict better response to SCC-DBS in TRD patients. We measured rACC glutamate and glutamine using pre-operative in vivo proton MRS (1H-MRS) and related levels to SCC-DBS response at 6 months in 16 TRD patients. Patients with bipolar disorder have demonstrated primarily opposite trends for diseaserelated glutamatergic alterations in the rACC, meaning increased Glx (Dager et al., 2004; Frye et al., 2007; Patel et al., 2008) compared to MDD. Therefore, we further investigated the impact of comorbid history of BPD on rACC biomarkers in our patient sample as an exploratory analysis.
2. Methods 2.1. Participants Sixteen patients with chronic treatment-resistant depression were included. Patients were part of a larger clinical trial, including 22 patients; results reported elsewhere (Ramasubbu et al., 2020). Due to technical restrictions, 1H-MRS data could not be collected in 6 patients. All patients met the DSM-IV/V requirements for MDD and were also deemed treatment-resistant (Fava, 2003). Among 16 patients included in this study, 2 had bipolar depression (BD). These patients were not differentiated from MDD cohort in the main study but were considered separately as an exploratory analysis in the current work. The University of Calgary Ethics Review Board approved the protocol and all patients gave informed consent (Clinicaltrials.gov NCT01983904).
2.2. DBS surgery and treatment protocol
Patients received bilateral SCC-DBS implantation. Details of clinical measurements, patient selection and stimulation protocols are described in our clinical outcome paper (Ramasubbu et al., 2020). Quadripolar DBS electrodes (3387; Medtronic, Minneapolis MN) were implanted such that at least 1–2 poles of the DBS lead were in the SCC target. The DBS electrodes were later connected to the subcutaneously implanted pulse generator (Activa PC; Medtronic) and stimulation was turned on one week after discharge. Monopolar stimulation at 130 Hz was applied with either pulse width (90– 450 μs), or amplitude (4–8 V) progressively increased monthly based on response status.
2.3. Clinical rating Clinical outcome was completed by the study psychiatrist, RR, using the clinician-rated Hamilton Depression Rating Scale 17 item (HDRS) (HAMILTON, 1960). Response was defined as a 50% reduction in HDRS from baseline to 6 months. Details are available in the main clinical manuscript (Ramasubbu et al., 2020).
2.4. Magnetic resonance imaging MRI data were acquired within a month of surgery, in a single session. T1-weighted and MRS data were acquired on a 3T General Electric Discovery 750 using a 12channel phased-array head coil. High-resolution T1 weighted images were collected using a 3D magnetization-prepared rapid gradient-echo (MP-RAGE) sequence with the following parameters: TE = 2.5ms, TR = 7.5ms, TI = 650ms; a flip angle of 11°, voxel size 0.9x0.9x0.9mm; number of slices = 236 (thickness 0.9 mm) and used for placing the MRS voxel. In order to investigate whether alterations in rACC grey matter volume related to glutamate levels, T1 brain images were automatically segmented using freely available software FreeSurfer v11.4.2 (http://surfer.nmr.mgh.harvard.edu/) to extract the volume of the rACC in each patient.
2.5. 1H-MRS Short echo (TE/TR: 35/1500 ms, 128 averages) PRESS proton spectroscopy was conducted with a 10x35x20mm (7ml) voxel placed in the pregenual anterior cingulate gyrus (Fig. 1A). The voxel was placed in the anterior cingulate gyrus, anterior to the genu of the corpus callosum as determined on a midsagittal slice. The anterior cingulate gyrus voxel contained both gray and white matter, primarily the rostral anterior cingulate gyrus. Voxel was carefully placed by experienced members of the research team, following illustrations (Fig. 1A) and detailed instructions to ensure consistency across participants. LCModel (Provencher, 2001) was used to calculate absolute metabolite concentrations (mmol/kg ww) for: glutamate and glutamine. MRS has been shown to have excellent test-retest reliability for such measures (Yasen et al., 2017). Cramer–Rao bounds were ≤12 for glutamate. Glutamine is more difficult to fit, so we used a more liberal cut-off of 30, as done previously (Lebel et al., 2016), which eliminated 3 individuals from the analysis. Figure 1 shows representative spectra for a responder and a non-responder, including the isolated spectra for glutamate and glutamine.
2.6. Statistics Metabolite levels at baseline were compared between responders and non-responders. Data were non-normal in distribution, therefore non-parametric Mann-Whitney U testing was used to examine the significance. Partial correlation, using age as a covariate of non-interest (Pearson R; one-tailed) determined whether baseline rACC glutamate or glutamine predicted symptom improvement. Significance was set at pvalue of 0.05, modified to 0.025, after Bonferroni correction for 2 metabolites. Exploratory analyses were performed excluding BD.
2.7. Data Availability Data will be made available upon request.
3. Results
3.1. Demographics Table 1 showed baseline characteristics of entire sample (n=22) involved in the DBS clinical trial (Ramasubbu et al., 2020) and the patients (n=16) included in this MRS study. Among those this study, 6 were classified as responders (MDD-4; BD-2) and 10 (MDD-10) as non-responders. Responders and non-responders included in this MRS study, did not significantly differ in age, gender, baseline HRSD score, duration of illness, number of episodes or number of medications. Mean age of participants was 47.4 with a range of 22-69 years old. HDRS was significantly reduced 6 months postDBS from baseline (F=19.841; p=0.002; Table 1).
3.2. Glutamate Baseline glutamate levels did not relate to baseline HRSD score. Baseline rACC glutamate levels related to 6-month change in HDRS scores (r=-0.561, p=0.018; Fig2B), but Mann-Whitney U analysis showed no categorical difference between responder and non-responder groups (p=0.13; Fig2A). Response groups became significantly difference with the exclusion of the 2 BD patients (p=0.013) and the relationship between rACC glutamate and symptom change was strengthened (r=-0.748, p=0.002).
3.3. Glutamine Baseline glutamine levels did not relate to baseline HDRS score. Baseline rACC glutamine levels also did not predict symptom change after 6 months (r=0.070, p=0.419) and there was no difference between responders and non-responders. Exclusion of patients with BD did not alter results.
3.4. Glutamate/Glutamine ratio The ratio of glutamate to glutamine levels also did not relate to baseline HDRS score, nor did it predict symptom change at 6 months. Exclusion of patients with BD did not alter results.
3.5. Volumetric analysis of rACC The volume of the rACC did not correlate with glutamate (r=0.292; p=0.172) or glutamine (r=-0.009; p=0.977) levels.
4. Discussion This is the first study to our knowledge investigating whether MRS-measured brain metabolites predicted response in patients undergoing SCC-DBS for TRD. We show lower rACC glutamate at the pre-treatment baseline predicted greater symptom improvement at 6-months post-DBS. This finding was unrelated to the effects of age or the volume of the rACC. Furthermore, exclusion of patients with bipolar treatment resistant depression from the analysis, increased the significance of the relationship between symptom reduction and rACC glutamate. Our results suggest that rACCglutamate could serve as a predictive biomarker of SCC-DBS outcomes, particularly in patients with treatment resistant major depression. MRS-based imaging would be an ideal DBS biomarker, as it is non-invasive, non-ionizing, and easily appended to standard preoperative MRI scans. We hypothesize that lower rACC glutamate levels in our patient responder population indicated pathology specifically involving the circuitry of the anterior cingulate region which may have improved of targeting an SCC circuit hub, with DBS. Both lower rACC and higher SCC metabolism have been linked to depressive pathology (Mayberg et al., 1997; Pizzagalli, 2011). We recently demonstrated that greater SCC metabolic activity (FDG-PET) also predicts 6-month DBS-response (Brown et al., 2018), in this cohort. Therefore, our combined data suggests decreased rACC and increased SCC metabolism in the target circuit biases toward better SCC-DBS response. However, the opposite appears true for response to antidepressants or placebo in nonresistant MDD (Mayberg et al., 1997; Pizzagalli, 2011). This differential response prediction of ACC circuit alterations requires further study.
It is also possible that glutamate levels impacted the efficacy of DBS, directly. Animals models demonstrate that the effect of DBS can be modulated by local glutamate level (Ross et al., 2016) and that blockade of glutamatergic transmission prevents the DBS-evoked LFP responses at distal sites (McCracken and Grace, 2007). High frequency stimulation of SCC may activate efferent axons projecting to rACC improving excitatory neural signalling in rACC, potentially correcting aberrant functioning in default mode, and salience networks. Glutamine levels in rACC did not predict DBS response. Considering glutamine is a marker of glial activity, it is possible that glial activity in rACC has a limited role in DBS response. Rostral ACC Glx concentrations have been proposed as a differential disease biomarker for BD and MDD (Lener et al., 2017; Taylor, 2014). Our data are consistent with that, as 2 responders with BD had higher pre-DBS rACC glutamate (10.09 and 9.16) level than the mean of sample 8.44±1.58, while we observed lower rACC glutamate in MDD responders.
4.1. Limitations Despite demonstrating a highly significant relationship between rACC glutamate and response, our sample size was small, so conclusions must be drawn with caution. The confidence with which we can report negative findings is especially limited. This is particularly relevant with respect to glutamine where lower brain levels increase measurement error. Furthermore, the positive findings of this study need replication and prospective validation in large independent sample to determine the predictive potentials of low glutamate level at rACC for patient selection in the future DBS trials. Also, due to scan time constraints, we only measured from one voxel in the brain and cannot ascertain whether the response-related glutamate alterations are specific to the rACC. The use of antidepressants or psychotropic medications may alter glutamate levels; however, both groups were equally exposed to multiple trials of antidepressants prior to DBS and therefore the differences in glutamate levels between groups cannot be explained by antidepressant treatment. We did not have healthy normative data for
comparison, so it was not possible to determine whether abnormalities in rACC glutamate levels was a marker of depression illness in addition to DBS response state.
4.2. Conclusions Overall, we demonstrate evidence that rACC glutamate could serve as a response biomarker for SCC-DBS in patients with TRD. Further, our exploratory analysis suggests that patient history of BD lowers the predictive value of this marker, but more work in a larger sample is needed to confirm this preliminary finding. Our data underscore the importance of pre-existing brain states to outcome in DBS and support the need for biomarker-based patient selection strategies to optimize response to this promising, yet invasive therapy. If our findings are replicated, MRS-measured glutamate levels in the rACC could form the basis for such a selection strategy for TRD patients being screened for SCC-DBS.
Contributors RR and ZHK were the principle investigators of the study. RR designed this study and performed all clinical evaluations and ZHK performed the DBS surgeries. DLC and ECB collected the data. DLC ran the analyses and prepared the first draft of the paper. FPM helped with data acquisition and analysis strategies. All authors edited manuscript drafts. All authors commented on earlier versions and approved the final manuscript.
Acknowledgements The authors would like to thank all the participants and their families. We also thank Bradley Goodyear and Filomeno Cortese for assistance with imaging data acquisition and handling.
Funding This study was supported by Alberta Innovates – Health Solutions Collaborative Research and Innovation Opportunity (AIHS-CRIO) Grant. DLC and ECB were supported by an AIHS Postdoctoral Fellowship. Preliminary results for this study were presented at Society of Biological Psychiatry (SOBP) 71st Meeting in Atlanta, Georgia, USA, 2016.
Competing Interests Dr. Ramasubbu has received honorarium for serving in the advisory committee of Astra Zeneca, Lundbeck, Janssen, and Otsuka. He also received investigator-initiated grant from Astra Zeneca and Pfizer. Other authors declare no potential conflict of interest. Dr. Kiss reports that Alberta health services receives value adds for research and education from Medtronic Canada, the manufacturer of the devices used in the study. These funds are only partially available for research to the authors for research and education through the Neuromodulation program. Medtronic had no involvement in any of this study.
Contributors RR and ZHK were the principle investigators of the study. RR designed this study and performed all clinical evaluations and ZHK performed the DBS surgeries. DC and ECB collected the data. DC ran the analyses and prepared the first draft of the paper. FPM helped with data acquisition and analysis strategies. All authors edited manuscript drafts. All authors commented on earlier versions and approved the final manuscript.
Acknowledgements The authors would like to thank all the participants and their families. We also thank Bradley Goodyear and Filomeno Cortese for assistance with imaging data acquisition and handling.
Funding This study was supported by Alberta Innovates – Health Solutions Collaborative Research and Innovation Opportunity (AIHS-CRIO) Grant. DLC and ECB were supported by an AIHS Postdoctoral Fellowship. Preliminary results for this study were presented at Society of Biological Psychiatry (SOBP) 71st Meeting in Atlanta, Georgia, USA, 2016.
The University of Calgary Ethics Review Board approved the protocol and all patients gave informed consent (Clinicaltrials.gov NCT01983904).
Competing Interests Dr. Ramasubbu has received honorarium for serving in the advisory committee of Astra Zeneca, Lundbeck, Janssen, and Otsuka. He also received investigator-initiated grant from Astra Zeneca and Pfizer. Other authors declare no potential conflict of interest. Dr. Kiss reports that Alberta health services receives value adds for research and education from Medtronic Canada, the manufacturer of the devices used in the study. These funds are only partially available for research to the authors for research and
education through the Neuromodulation program. Medtronic had no involvement in any of this study.
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Figure 1: Location of the rACC MRS voxel with respect to the approximate position of the SCC-DBS electrode (A). The LCModel fit in a DBS responder (7017; B) and nonresponder (7011; C) for glutamate and glutamine.
Full Clinical Sample (n=22)
Age Male/Female Duration of illness (years) Duration of current episode (months) Baseline HDRS
Current Study Sample (n=16)
Responders (N=10)
Non-responders Responders (N=12) (N=6)
39.1 ±3.8 4/6
52.5 ±4.0 * 8/4
41.0 ±6.09 1/5
Nonresponders (N=10) 51.3 ±4.6 6/4
18.9 ±3.6
28.2 ±5.4
21.0 ±8.32
25.1 ±5.6
23.6 ±7.2 24.2 ±1.27
24.3 ±4.7 23.3 ±1.11
25.5 ±10.94 23.3 ±1.33
28.6 ±6.75 23.5 ±1.28
19.0 ±1.45 #
Note: HDRS = Hamilton Depression Rating Scale (17 item); *p<0.05 **p<0.001 significant difference between responders and non-responders of full clinical sample. #p<0.005 significant difference between responders and non-responders of current study sample. 6-Month HDRS
8.00 ±1.33
18.3 ±1.30 **
9.0 ±1.69
Table 1: Demographic features of responders and non-responders in the full clinical sample (adapted from Ramasubbu et al., 2020) and the sample included in this biomarker study. Data presented as mean ±SEM.
Figure 2: A) Glutamate levels are significantly lower in responders, compared to non-responders in MDD patients, excluding BD patients and B) rACC glutamate negatively correlates with the change in HDRS at 6 months.
Rostral anterior cingulate glutamate predicts response to subcallosal deep brain stimulation for resistant depression Darren L. Clark , Frank P. MacMaster , Elliot C. Brown , Zelma H.T. Kiss , Rajamannar Ramasubbu PII: DOI: Reference:
S0165-0327(19)33129-5 https://doi.org/10.1016/j.jad.2020.01.058 JAD 11507
To appear in:
Journal of Affective Disorders
Received date: Revised date: Accepted date:
8 November 2019 9 January 2020 14 January 2020
Please cite this article as: Darren L. Clark , Frank P. MacMaster , Elliot C. Brown , Zelma H.T. Kiss , Rajamannar Ramasubbu , Rostral anterior cingulate glutamate predicts response to subcallosal deep brain stimulation for resistant depression, Journal of Affective Disorders (2020), doi: https://doi.org/10.1016/j.jad.2020.01.058
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Highlights Sham-controlled SCC-DBS trial showed negative results in patients with TRD Biomarker-stratified patient selection may improve the SCC-DBS outcomes Lower baseline rACC-Glu (1H -MRS) predicted response at 6 months in 16 TRD patients Excluding comorbid bipolar disorder patients improved prediction Pre-DBS rACC-Glu level may have potentials as a biomarker for patient selection
Rostral anterior cingulate glutamate predicts response to subcallosal deep brain stimulation for resistant depression
Darren L. Clark1-4, Frank P. MacMaster1,3-6, Elliot C. Brown1-4, Zelma H. T. Kiss2-4, Rajamannar Ramasubbu1,3,4. 1
Department of Psychiatry, University of Calgary, Calgary, AB, Canada
2 3
Department of Clinical Neuroscience, University of Calgary, AB, Canada
Mathison Centre for Mental Health Research and Education, University of Calgary,
Calgary, AB, Canada 4
Hotchkiss Brain Institute, University of Calgary, Calgary, AB, Canada
5
Department of Radiology, University of Calgary, Calgary, AB, Canada
6
Child and Adolescent Imaging Research Program, Alberta Children’s Hospital, Calgary,
AB, Canada
*CORRESPONDENCE: Rajamannar Ramasubbu MD, Department of Psychiatry/ Clinical Neurosciences, University of Calgary, Mathison Centre for Mental Health Research and Education, TRW building, Room 4D64, 3280 Hospital Drive NW, Calgary, Alberta, T2N4Z6 Tel +1 403 210 6890; Fax +1 403 210 9114, Email: [email protected].
Keywords: deep brain stimulation, treatment-resistant depression, glutamate, glutamine, anterior cingulate cortex, magnetic resonance imaging
Abstract Background: Deep brain stimulation (DBS) of the subcallosal cingulate (SCC) provided benefit for treatment-resistant depression (TRD) in open-label studies but failed in a
recent randomized sham-controlled trial. Informed patient selection, based on reliable biomarkers, is needed to optimize outcome. We investigated if rostral anterior cingulate (rACC) glutamate/glutamine concentration could serve as a potential biomarker of response. Methods: Sixteen adults with TRD (Major Depression; MDD = 14; Bipolar Depression; BD =2) underwent proton magnetic resonance spectroscopy using a short-echo proton spectroscopy with a voxel placed in the rACC, prior to DBS. Improvement in depression was assessed using the 17-item Hamilton Rating Scale for Depression (HRSD). Glutamate and glutamine concentrations at baseline in the rACC were examined in relation to clinical outcomes at six months. Results: Lower baseline glutamate predicted significant reduction in HRSD scores in all patients (p=0.018), and predicted both HDRS reduction (p=0.002) and 6-month response outcome in MDD patients (p=0.013). Neither baseline glutamine, nor glutamine/glutamate ratio significantly related to outcome nor symptom improvement. Limitations: Our study was limited by sample size, though it is large for a DBS study. We measured from a single voxel in the brain, so we cannot be certain our findings are specific to the rACC. Conclusions: These preliminary results suggest that baseline rACC-glutamate concentration could serve as a response-predictive biomarker for SCC-DBS, particularly in patients with resistant major depression. If our findings are replicated and validated, rACC-glutamate may provide a basis to prospectively select TRD patients to improve likelihood of response to SCC-DBS.
1. Introduction Subcallosal cingulate (SCC) deep brain stimulation (DBS) is a promising investigational intervention for treatment-resistant depression (TRD) with 6-month response rates around 50% in open-label trials (Dandekar et al., 2018). Unfortunately, a double-blind randomized, controlled trial of SCC-DBS failed to demonstrate difference between sham and active DBS at 6-months (Holtzheimer et al., 2017). This failure has been
attributed to various factors, including inconsistent patient selection. DBS is invasive, costly and associated with significant risk. Informed patient selection strategies, based on reliable response biomarkers, would defray costs and limit the risks of invasive surgery to those most likely to benefit. The rostral anterior cingulate (rACC; Brodmann area (BA)24) lies anterior and dorsal to the SCC target region and has emerged as a promising biomarker of illness and predictor of treatment response in depression (reviewed in Pizzagalli, 2011 (Pizzagalli, 2011)). Altered glutamatergic signaling in the rACC is implicated in MDD pathology. Reduced glutamate, glutamine or combined glutamate-glutamine (Glx) levels, measured with proton magnetic resonance spectroscopy (MRS), have been described in the rACC of adults (Auer et al., 2000; Merkl et al., 2011) and youths (Mirza et al., 2004) with MDD. Lower rACC glutamate levels have been related to anhedonia, both directly through relationship with subjective emotional rating and indirectly via correlation with reduced negative bold response to emotional stimuli (Walter et al., 2009). It was also related to reduced resting functional connectivity of the rACC to the anterior insula, a network required for proper saliency processing and switching between default-mode network and executive function (Horn et al., 2010). Altered glutamate levels in the rACC might represent a failure of the greater ACC circuit to properly regulate emotional responses, and facilitate task-based network switching, contributing to emotional dysregulation and perseverative rumination. Furthermore, The rACC is tightly interconnected to the SCC (Heilbronner and Haber, 2014; Palomero-Gallagher et al., 2008) and the function of these regions is highly (though often inversely) correlated (Chiba et al., 2001; Johansen-Berg et al., 2008; Pizzagalli, 2011). Stimulation of the local SCC with DBS would likely alter the broader ACC function, either directly via cingulate white matter activation (Riva-Posse et al., 2018; 2014) or indirectly via local SCC activity changes impacting the function of rACC via reciprocal connections (Chiba et al., 2001; Heilbronner and Haber, 2014; Johansen-Berg et al., 2008). Although direct evidence Is lacking, preclinical studies suggest that DBS may modulate excitatory glutamatergic neurotransmission (Ross et al., 2016). In addition, pre-treatment glutamatergic levels in
the rACC predicted response and/or were normalized after electroconvulsive therapy (ECT) in patients with TRD (Pfleiderer et al., 2003; Zhang et al., 2013), providing further support for the potential utility of rACC glutamatergic signaling as a predictor and response biomarker for SCC-DBS in TRD patients. We hypothesized that lower rACC glutamate or glutamine levels would predict better response to SCC-DBS in TRD patients. We measured rACC glutamate and glutamine using pre-operative in vivo proton MRS (1H-MRS) and related levels to SCC-DBS response at 6 months in 16 TRD patients. Patients with bipolar disorder have demonstrated primarily opposite trends for diseaserelated glutamatergic alterations in the rACC, meaning increased Glx (Dager et al., 2004; Frye et al., 2007; Patel et al., 2008) compared to MDD. Therefore, we further investigated the impact of comorbid history of BPD on rACC biomarkers in our patient sample as an exploratory analysis.
2. Methods 2.1. Participants Sixteen patients with chronic treatment-resistant depression were included. Patients were part of a larger clinical trial, including 22 patients; results reported elsewhere (Ramasubbu et al., 2020). Due to technical restrictions, 1H-MRS data could not be collected in 6 patients. All patients met the DSM-IV/V requirements for MDD and were also deemed treatment-resistant (Fava, 2003). Among 16 patients included in this study, 2 had bipolar depression (BD). These patients were not differentiated from MDD cohort in the main study but were considered separately as an exploratory analysis in the current work. The University of Calgary Ethics Review Board approved the protocol and all patients gave informed consent (Clinicaltrials.gov NCT01983904).
2.2. DBS surgery and treatment protocol
Patients received bilateral SCC-DBS implantation. Details of clinical measurements, patient selection and stimulation protocols are described in our clinical outcome paper (Ramasubbu et al., 2020). Quadripolar DBS electrodes (3387; Medtronic, Minneapolis MN) were implanted such that at least 1–2 poles of the DBS lead were in the SCC target. The DBS electrodes were later connected to the subcutaneously implanted pulse generator (Activa PC; Medtronic) and stimulation was turned on one week after discharge. Monopolar stimulation at 130 Hz was applied with either pulse width (90– 450 μs), or amplitude (4–8 V) progressively increased monthly based on response status.
2.3. Clinical rating Clinical outcome was completed by the study psychiatrist, RR, using the clinician-rated Hamilton Depression Rating Scale 17 item (HDRS) (HAMILTON, 1960). Response was defined as a 50% reduction in HDRS from baseline to 6 months. Details are available in the main clinical manuscript (Ramasubbu et al., 2020).
2.4. Magnetic resonance imaging MRI data were acquired within a month of surgery, in a single session. T1-weighted and MRS data were acquired on a 3T General Electric Discovery 750 using a 12channel phased-array head coil. High-resolution T1 weighted images were collected using a 3D magnetization-prepared rapid gradient-echo (MP-RAGE) sequence with the following parameters: TE = 2.5ms, TR = 7.5ms, TI = 650ms; a flip angle of 11°, voxel size 0.9x0.9x0.9mm; number of slices = 236 (thickness 0.9 mm) and used for placing the MRS voxel. In order to investigate whether alterations in rACC grey matter volume related to glutamate levels, T1 brain images were automatically segmented using freely available software FreeSurfer v11.4.2 (http://surfer.nmr.mgh.harvard.edu/) to extract the volume of the rACC in each patient.
2.5. 1H-MRS Short echo (TE/TR: 35/1500 ms, 128 averages) PRESS proton spectroscopy was conducted with a 10x35x20mm (7ml) voxel placed in the pregenual anterior cingulate gyrus (Fig. 1A). The voxel was placed in the anterior cingulate gyrus, anterior to the genu of the corpus callosum as determined on a midsagittal slice. The anterior cingulate gyrus voxel contained both gray and white matter, primarily the rostral anterior cingulate gyrus. Voxel was carefully placed by experienced members of the research team, following illustrations (Fig. 1A) and detailed instructions to ensure consistency across participants. LCModel (Provencher, 2001) was used to calculate absolute metabolite concentrations (mmol/kg ww) for: glutamate and glutamine. MRS has been shown to have excellent test-retest reliability for such measures (Yasen et al., 2017). Cramer–Rao bounds were ≤12 for glutamate. Glutamine is more difficult to fit, so we used a more liberal cut-off of 30, as done previously (Lebel et al., 2016), which eliminated 3 individuals from the analysis. Figure 1 shows representative spectra for a responder and a non-responder, including the isolated spectra for glutamate and glutamine.
2.6. Statistics Metabolite levels at baseline were compared between responders and non-responders. Data were non-normal in distribution, therefore non-parametric Mann-Whitney U testing was used to examine the significance. Partial correlation, using age as a covariate of non-interest (Pearson R; one-tailed) determined whether baseline rACC glutamate or glutamine predicted symptom improvement. Significance was set at pvalue of 0.05, modified to 0.025, after Bonferroni correction for 2 metabolites. Exploratory analyses were performed excluding BD.
2.7. Data Availability Data will be made available upon request.
3. Results
3.1. Demographics Table 1 showed baseline characteristics of entire sample (n=22) involved in the DBS clinical trial (Ramasubbu et al., 2020) and the patients (n=16) included in this MRS study. Among those this study, 6 were classified as responders (MDD-4; BD-2) and 10 (MDD-10) as non-responders. Responders and non-responders included in this MRS study, did not significantly differ in age, gender, baseline HRSD score, duration of illness, number of episodes or number of medications. Mean age of participants was 47.4 with a range of 22-69 years old. HDRS was significantly reduced 6 months postDBS from baseline (F=19.841; p=0.002; Table 1).
3.2. Glutamate Baseline glutamate levels did not relate to baseline HRSD score. Baseline rACC glutamate levels related to 6-month change in HDRS scores (r=-0.561, p=0.018; Fig2B), but Mann-Whitney U analysis showed no categorical difference between responder and non-responder groups (p=0.13; Fig2A). Response groups became significantly difference with the exclusion of the 2 BD patients (p=0.013) and the relationship between rACC glutamate and symptom change was strengthened (r=-0.748, p=0.002).
3.3. Glutamine Baseline glutamine levels did not relate to baseline HDRS score. Baseline rACC glutamine levels also did not predict symptom change after 6 months (r=0.070, p=0.419) and there was no difference between responders and non-responders. Exclusion of patients with BD did not alter results.
3.4. Glutamate/Glutamine ratio The ratio of glutamate to glutamine levels also did not relate to baseline HDRS score, nor did it predict symptom change at 6 months. Exclusion of patients with BD did not alter results.
3.5. Volumetric analysis of rACC The volume of the rACC did not correlate with glutamate (r=0.292; p=0.172) or glutamine (r=-0.009; p=0.977) levels.
4. Discussion This is the first study to our knowledge investigating whether MRS-measured brain metabolites predicted response in patients undergoing SCC-DBS for TRD. We show lower rACC glutamate at the pre-treatment baseline predicted greater symptom improvement at 6-months post-DBS. This finding was unrelated to the effects of age or the volume of the rACC. Furthermore, exclusion of patients with bipolar treatment resistant depression from the analysis, increased the significance of the relationship between symptom reduction and rACC glutamate. Our results suggest that rACCglutamate could serve as a predictive biomarker of SCC-DBS outcomes, particularly in patients with treatment resistant major depression. MRS-based imaging would be an ideal DBS biomarker, as it is non-invasive, non-ionizing, and easily appended to standard preoperative MRI scans. We hypothesize that lower rACC glutamate levels in our patient responder population indicated pathology specifically involving the circuitry of the anterior cingulate region which may have improved of targeting an SCC circuit hub, with DBS. Both lower rACC and higher SCC metabolism have been linked to depressive pathology (Mayberg et al., 1997; Pizzagalli, 2011). We recently demonstrated that greater SCC metabolic activity (FDG-PET) also predicts 6-month DBS-response (Brown et al., 2018), in this cohort. Therefore, our combined data suggests decreased rACC and increased SCC metabolism in the target circuit biases toward better SCC-DBS response. However, the opposite appears true for response to antidepressants or placebo in nonresistant MDD (Mayberg et al., 1997; Pizzagalli, 2011). This differential response prediction of ACC circuit alterations requires further study.
It is also possible that glutamate levels impacted the efficacy of DBS, directly. Animals models demonstrate that the effect of DBS can be modulated by local glutamate level (Ross et al., 2016) and that blockade of glutamatergic transmission prevents the DBS-evoked LFP responses at distal sites (McCracken and Grace, 2007). High frequency stimulation of SCC may activate efferent axons projecting to rACC improving excitatory neural signalling in rACC, potentially correcting aberrant functioning in default mode, and salience networks. Glutamine levels in rACC did not predict DBS response. Considering glutamine is a marker of glial activity, it is possible that glial activity in rACC has a limited role in DBS response. Rostral ACC Glx concentrations have been proposed as a differential disease biomarker for BD and MDD (Lener et al., 2017; Taylor, 2014). Our data are consistent with that, as 2 responders with BD had higher pre-DBS rACC glutamate (10.09 and 9.16) level than the mean of sample 8.44±1.58, while we observed lower rACC glutamate in MDD responders.
4.1. Limitations Despite demonstrating a highly significant relationship between rACC glutamate and response, our sample size was small, so conclusions must be drawn with caution. The confidence with which we can report negative findings is especially limited. This is particularly relevant with respect to glutamine where lower brain levels increase measurement error. Furthermore, the positive findings of this study need replication and prospective validation in large independent sample to determine the predictive potentials of low glutamate level at rACC for patient selection in the future DBS trials. Also, due to scan time constraints, we only measured from one voxel in the brain and cannot ascertain whether the response-related glutamate alterations are specific to the rACC. The use of antidepressants or psychotropic medications may alter glutamate levels; however, both groups were equally exposed to multiple trials of antidepressants prior to DBS and therefore the differences in glutamate levels between groups cannot be explained by antidepressant treatment. We did not have healthy normative data for
comparison, so it was not possible to determine whether abnormalities in rACC glutamate levels was a marker of depression illness in addition to DBS response state.
4.2. Conclusions Overall, we demonstrate evidence that rACC glutamate could serve as a response biomarker for SCC-DBS in patients with TRD. Further, our exploratory analysis suggests that patient history of BD lowers the predictive value of this marker, but more work in a larger sample is needed to confirm this preliminary finding. Our data underscore the importance of pre-existing brain states to outcome in DBS and support the need for biomarker-based patient selection strategies to optimize response to this promising, yet invasive therapy. If our findings are replicated, MRS-measured glutamate levels in the rACC could form the basis for such a selection strategy for TRD patients being screened for SCC-DBS.
Contributors RR and ZHK were the principle investigators of the study. RR designed this study and performed all clinical evaluations and ZHK performed the DBS surgeries. DLC and ECB collected the data. DLC ran the analyses and prepared the first draft of the paper. FPM helped with data acquisition and analysis strategies. All authors edited manuscript drafts. All authors commented on earlier versions and approved the final manuscript.
Acknowledgements The authors would like to thank all the participants and their families. We also thank Bradley Goodyear and Filomeno Cortese for assistance with imaging data acquisition and handling.
Funding This study was supported by Alberta Innovates – Health Solutions Collaborative Research and Innovation Opportunity (AIHS-CRIO) Grant. DLC and ECB were supported by an AIHS Postdoctoral Fellowship. Preliminary results for this study were presented at Society of Biological Psychiatry (SOBP) 71st Meeting in Atlanta, Georgia, USA, 2016.
Competing Interests Dr. Ramasubbu has received honorarium for serving in the advisory committee of Astra Zeneca, Lundbeck, Janssen, and Otsuka. He also received investigator-initiated grant from Astra Zeneca and Pfizer. Other authors declare no potential conflict of interest. Dr. Kiss reports that Alberta health services receives value adds for research and education from Medtronic Canada, the manufacturer of the devices used in the study. These funds are only partially available for research to the authors for research and education through the Neuromodulation program. Medtronic had no involvement in any of this study.
Contributors RR and ZHK were the principle investigators of the study. RR designed this study and performed all clinical evaluations and ZHK performed the DBS surgeries. DC and ECB collected the data. DC ran the analyses and prepared the first draft of the paper. FPM helped with data acquisition and analysis strategies. All authors edited manuscript drafts. All authors commented on earlier versions and approved the final manuscript.
Acknowledgements The authors would like to thank all the participants and their families. We also thank Bradley Goodyear and Filomeno Cortese for assistance with imaging data acquisition and handling.
Funding This study was supported by Alberta Innovates – Health Solutions Collaborative Research and Innovation Opportunity (AIHS-CRIO) Grant. DLC and ECB were supported by an AIHS Postdoctoral Fellowship. Preliminary results for this study were presented at Society of Biological Psychiatry (SOBP) 71st Meeting in Atlanta, Georgia, USA, 2016.
The University of Calgary Ethics Review Board approved the protocol and all patients gave informed consent (Clinicaltrials.gov NCT01983904).
Competing Interests Dr. Ramasubbu has received honorarium for serving in the advisory committee of Astra Zeneca, Lundbeck, Janssen, and Otsuka. He also received investigator-initiated grant from Astra Zeneca and Pfizer. Other authors declare no potential conflict of interest. Dr. Kiss reports that Alberta health services receives value adds for research and education from Medtronic Canada, the manufacturer of the devices used in the study. These funds are only partially available for research to the authors for research and
education through the Neuromodulation program. Medtronic had no involvement in any of this study.
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Figure 1: Location of the rACC MRS voxel with respect to the approximate position of the SCC-DBS electrode (A). The LCModel fit in a DBS responder (7017; B) and nonresponder (7011; C) for glutamate and glutamine.
Full Clinical Sample (n=22)
Age Male/Female Duration of illness (years) Duration of current episode (months) Baseline HDRS
Current Study Sample (n=16)
Responders (N=10)
Non-responders Responders (N=12) (N=6)
39.1 ±3.8 4/6
52.5 ±4.0 * 8/4
41.0 ±6.09 1/5
Nonresponders (N=10) 51.3 ±4.6 6/4
18.9 ±3.6
28.2 ±5.4
21.0 ±8.32
25.1 ±5.6
23.6 ±7.2 24.2 ±1.27
24.3 ±4.7 23.3 ±1.11
25.5 ±10.94 23.3 ±1.33
28.6 ±6.75 23.5 ±1.28
19.0 ±1.45 #
Note: HDRS = Hamilton Depression Rating Scale (17 item); *p<0.05 **p<0.001 significant difference between responders and non-responders of full clinical sample. #p<0.005 significant difference between responders and non-responders of current study sample. 6-Month HDRS
8.00 ±1.33
18.3 ±1.30 **
9.0 ±1.69
Table 1: Demographic features of responders and non-responders in the full clinical sample (adapted from Ramasubbu et al., 2020) and the sample included in this biomarker study. Data presented as mean ±SEM.
Figure 2: A) Glutamate levels are significantly lower in responders, compared to non-responders in MDD patients, excluding BD patients and B) rACC glutamate negatively correlates with the change in HDRS at 6 months.