Investigation of Poststroke Depression Following a Nucleus Accumbens Infarct in Mice

Abstract

BACKGROUND:

Poststroke depression (PSD) affects ≈33% of individuals 1 year after a stroke. Blood-brain barrier (BBB) dysfunction in the nucleus accumbens (NAc), a hub for emotional processing, reward, and mood regulation, has been linked to stress-induced depressive-like behaviors in male mice. Neurovascular alterations were also observed in postmortem tissue samples from men with a diagnosis of major depression. Thus, we aimed to investigate if BBB changes in the NAc could contribute to PSD pathophysiology.

METHODS:

Stereotaxic injection of ET-1 (endothelin-1), a potent vasoconstrictor, was performed in the NAc of male mice to create a focal brain stroke, and then, infarct size and localization were assessed and quantified. We subsequently evaluated transcriptomic and morphological effects of the infarct on BBB-related genes and cells in the NAc, particularly those known to be altered after stress exposure in mice or human depression. BBB integrity was assessed with a dextran dye, and magnetic resonance imaging scans were conducted before versus after the injection of Gadovist, a contrast agent. Last, a battery of behavioral tests related to depressive- and anxiety-like behaviors was performed to determine if an infarct in the NAc is sufficient to induce a PSD-like phenotype.

RESULTS:

Following ET-1 injection, ≈50% of the total lesion was observed in the NAc leading to BBB hyperpermeability in this brain area. BBB gene expression was impacted by ET-1, and also surgery alone and profiles were differentially regulated throughout time up to 14 days. Gliosis in the NAc was observed with increased reactivity of astrocytes and microglia. The effect of ET-1 on PSD-like symptoms was limited. However, body weight, sociability, and activity were affected by surgery with a more pronounced impact of ET-1 on social interactions compared with naive animals.

CONCLUSIONS:

While no clear PSD phenotype was observed following an ET-1–induced stroke in the NAc of male mice, our study shed light on the technical complexity of focal lesions in deep brain structures, an understudied phenomenon occurring in humans. We provide technical insights for the development of a mouse model of deep brain lesions, characterize its impact at molecular, cellular, and behavioral levels, and highlight the need to control for vascular alterations when performing stroke surgeries.

Stroke is a neurological deficit due to a lesion of the central nervous system¹ and the second leading cause of mortality worldwide.² Psychiatric complications are frequent,³ the most prevalent being poststroke depression (PSD)³: 33% after 1 year and 23% after 5 years⁴; ≈85% of individuals with PSD had not experienced major depressive disorder (MDD) prior.⁵ Most PSD clinical features are similar to MDD with depressed mood, loss of interest, and anhedonia reported. However, individuals with PSD often experience more sleep disturbance, vegetative symptoms, and social withdrawal.⁶ The first treatment choice for PSD is classical selective serotonin reuptake inhibitor antidepressants⁶; however, only 30% of depressed individuals achieve complete remission.⁷ In the context of stroke, depressive episodes are often recurrent and hard to treat, suggesting that causal biological mechanisms remain untreated.

Lacunar infarcts, responsible for 20% of ischemic stroke,⁸ occur when small, penetrating branches of cerebral vessels are occluded in deep brain regions.^9,10 Lesions affecting frontal or basal ganglia areas are associated with cognitive deficits and PSD.^11–16 In rodents, the gold standard procedure to induce stroke is the middle cerebral artery occlusion (MCAO) model.¹⁷ It induces lesions of variable volume depending on the duration of the occlusion,¹⁸ generally affecting most of the caudo-putamen region,¹⁹ with this large nonspecific injury not recapitulating lacunar infarcts. Thus, developing focal models of ischemic stroke is of high clinical interest to elucidate the cellular and molecular mechanisms underlying PSD. Here, we took advantage of ET-1 (endothelin-1), a vasoactive agent causing constriction of local arterioles when injected into the brain, leading to reduced blood flow and ischemia,²⁰ to induce stroke in a region-specific manner. This approach has been used to provoke focal infarcts in brain areas such as the cortex,^21–24 hippocampus,²⁵ dorsal striatum,²⁶ and substantia nigra²⁷ but never successfully in deep ventral striatum subregions such as the nucleus accumbens (NAc).

The NAc is a structure in the reward system involved in addiction, motivation, reward, locomotion, and eating behaviors.^28,29 This brain region is affected in MDD with altered neuronal activity evidenced by magnetic resonance imaging scans³⁰ and linked to stress-induced anhedonia and social deficits in mice.³¹ Exposure to chronic stress induces blood-brain barrier (BBB) hyperpermeability in the NAc of male mice, along with depressive-like behaviors, via loss of tight junction protein Cldn5 (claudin-5) allowing passage of circulating inflammation.³² Loss of NAc Cldn5 was confirmed in men with MDD.³² To date, no study has investigated if a stroke in the NAc could promote PSD pathogenesis or decipher underlying mechanisms.

The BBB, formed by endothelial cells, pericytes, and astrocytes, regulates exchanges between the blood and the central nervous system in physiological conditions to maintain brain homeostasis.³³ Following stroke-induced ischemia, BBB properties and functions are impaired in a biphasic pattern.³⁴ Preclinical studies have demonstrated that promoting BBB integrity following ischemic lesions reduces the severity and volume of the infarct.^35–38 This led us to hypothesize that lacunar NAc infarcts could cause PSD by favoring BBB disruption and, eventually, neuronal dysfunction in this mood-regulating brain area. Therefore, we first optimized an ischemic mouse model of stroke in the NAc and then characterized its impact on BBB permeability, gene expression patterns, morphology, and development of anxiety- and depressive-like behaviors. Our results show that ET-1 stereotaxic injection induces a small and transient infarct in the NAc of male mice, which was not completely restricted to this brain area. The lesion successfully induced BBB-associated transcriptional changes, loss of BBB integrity, and gliosis, leading to lower social interactions without affecting other behavioral domains.

Methods

Please refer to the Supplemental Methods for additional details. All mouse procedures were performed in accordance with the Canadian Council on Animal Care, the Université Laval Animal Care Committee (2022-1056, VRR-22-1056), and ARRIVE guidelines (Animal Research: Reporting of In Vivo Experiments).³⁹ The data that support the findings of this study are available from the corresponding author upon reasonable request.

Eight- to 10-week-old male C57BL/6 mice were purchased from Charles River Laboratories. Anesthetized mice were infused with 2.0 μL of ET-1 (1 μg/μL diluted in nuclease-free sterile water⁴⁰) or water only (sham) into the left NAc.^32,41 To assess the infarct volume, brain sections of PFA-perfused mice were sliced, washed in ethanol, and incubated in cresyl violet and then xylene before mounting. Volume (mm³) was calculated as Σ (area of infarct in each section)×distance between each section and defined by the absence or reduced Nissl coloring.

BBB leakiness was evaluated by retro-orbital injection of Alexa Fluor 488-dextran⁴¹ or magnetic resonance imaging scans prior to and after administration of the Gadovist contrast agent. T1-weight images were preprocessed and analyzed with Advanced Normalization Tools^42,43 to quantify intensity lateralization. For transcriptional profiling, brain punches were collected, and RNA was isolated and reversed transcribed to cDNA for quantitative polymerase chain reaction analysis with gene expression normalized to Gapdh.^32,41,44 Primer pairs are listed in Table S1. For Cldn5 immunohistochemistry, whole brains were fresh-frozen, sliced, sections postfixed in methanol, washed, blocked, incubated overnight with primary antibodies, then appropriated secondary antibodies before mounting. Glial activation was assessed with immunohistochemistry of sections stained with anti-Gfap (glial fibrillary acidic protein), anti-NeuN (neuronal nuclei), and anti-Iba-1 (ionized calcium-binding adapter molecule 1) antibodies. Finally, behavioral studies for anxiety- and depression-like behaviors were performed as described previously.^{32,41,44–46}

The sample size was based on previous publications^32,40,41,47 and behavioral tests performed with an automated tracking system or scoring done by blinded experimenters. GraphPad Prism software (version 10.3) was used for statistical analyses, and significance was set at P<0.05.

Results

ET-1 Injection Induces a Small and Transient Infarct in the Left NAc

In rodents, focal injection of ET-1 induces localized brain infarcts,^26,27 but, to our knowledge, this had never been performed prior in deep brain structures such as the NAc. Thus, we first sought to optimize the protocol by injecting ET-1 or sterile water (sham) stereotaxically into the left NAc of C57BL/6 male mice and then quantifying the infarct lesions at 1, 3, 7, and 14 days post-injection using cresyl violet staining (Figure 1A). The extent of tissue damage to the whole left hemisphere was determined from the needle track itself (Figure 1B). Total lesion volumes were higher in ET-1–injected mice compared with sham animals until 7 days post-surgery (Figure 1C). The proportion of total lesions within the NAc was estimated at ≈50% up until 14 days after surgery with 65.64% of the lesions still present at this timepoint (Figure 1C). These results indicate the presence of nonspecific infarcted tissue along the needle track. Nevertheless, the lesion was larger in ET-1 versus sham animals in the left NAc until 7 days poststroke (Figure 1D). After 14 days, recovery was observed with volume of the infarct representing 23% of the original NAc lesion in ET-1–injected mice (Figure 1D). Next, BBB permeability was evaluated using peripheral injection of a fluorescent-tagged 10-kDa dextran at 1, 3, 7, and 14 days poststroke.⁴¹ ET-1 injection caused loss of BBB integrity in the NAc and passage of the dextran in the brain parenchyma, while it was restricted to the needle track in sham animals at late timepoints (Figure 1E; Figure S1). We also assessed BBB permeability with a gadolinium contrast agent and magnetic resonance imaging scans.³² Higher Gadovist signal was detected in the NAc of ET-1–injected male mice compared with sham (Figure 1F). Overall, these results suggest that ET-1 stereotaxic injection in the NAc can induce a small transient stroke and BBB disturbance; however, it is not perfectly restricted to this brain area.

Figure 1.

Establishment of the ET-1 (endothelin-1) unilateral stroke model in the mouse nucleus accumbens (NAc). A, Coronal section and coordinates for stereotaxic injection of either ET-1 or water (sham) in the left NAc and experimental timeline. Schematic illustration of ET-1 lesions across 10 sections and analysis strategies for total (left hemisphere) or NAc lesion quantification. B, Representative images of cresyl violet staining showing damage induced in a sham (left) vs ET-1–injected animal (right) at day (D) 1, D3, D7, and D14 post-surgery. C, Total lesion volume (mm³) across time for ET-1 vs sham-injected animals (2-way ANOVA: stroke×time interaction: *P=0.0121; n=3–4/group; D1: P≤0.0001; D3: P≤0.0001; D7: P=0.0009; and D14: P=0.1056). Pie charts represent the percentage of total ET-1 injury located in the NAc at D1 (48.64%), D3 (51.10%), D7 (53.73%), and D14 (65.64%) post-surgery. D, NAc lesion volume (mm³) across time for ET-1 vs sham-injected animals (2-way ANOVA: stroke effect: ****P≤0.0001; time effect: **P=0.0085; n=3–4/group; D1: P=0.0005; D3: P=0.0001; D7: P=0.0040; and D14: P=0.2558). Pie charts represent the percentage of D1 lesions in the NAc after ET-1 injury at D3 (86.1%), D7 (70.1%), and D14 (23.0%) post-surgery. E, ET-1–induced blood-brain barrier leakiness in the NAc was confirmed by retro-orbital injection of a 10-kDa dextran dye tagged with Alexa Fluor 488. Endothelial cells were stained with the Podxl (podocalyxin) marker and 4′,6-diamidino-2-phenylindole (DAPI) for nuclei. F, T1-weight magnetic resonance imaging (MRI) signal in the NAc before vs after intravenous injection of the contrast agent Gadovist in sham vs ET-1–injected mice at D7 (2-way ANOVA: Gadovist effect: **P=0.0017; n=5/group) and D14 (2-way ANOVA: stroke effect: *P=0.0189; n=5/group). T1 signal in the NAc left (injected) hemisphere was normalized on the right intact hemisphere. Data represent mean±SEM. Two-way ANOVA followed by the Tuckey multiple comparison test was applied with ****P≤0.0001, ***P≤0.001, **P≤0.01, and *P≤0.05.

BBB-Associated Gene Expression Is Affected by Surgery in Both Sham and ET-1 Animals

Following stroke, important vascular remodeling occurs, and the BBB is strongly altered.³⁴ We assessed time-dependent transcriptional responses of genes associated with endothelial cells (Pecam1), tight junctions (Cldn5, Ocln, and Tjp1), astrocyte reactivity (Gfap), and angiogenesis (Vegfa) at 1, 3, 7, and 14 days post-surgery (Figure 2A). Expression levels were normalized to gene expression at 1 day post-surgery for sham (left) and ET-1–injected mice (right; Figure 2A). Overall, transcriptional profiles across time seem similar between groups (Figure 2A). However, a significant decrease in Pecam1 expression was observed in the NAc of sham mice from D3 to D14 with this loss only emerging at D7 for ET-1–injected animals. Cldn5 and Ocln expressions are upregulated in ET-1–injected mice 7 and 14 days post-surgery, respectively, suggesting that tight junctions and BBB integrity are modulated by ET-1–induced infarct but not surgery itself (sham). The biphasic response of Tjp1 was almost identical in both groups pointing to a general surgery effect for this tight junction. Expression of Gfap, a marker for activated astrocytes, increased only in ET-1–injected mice at 7 days post-surgery, suggesting astrogliosis characteristic of stroke.⁴⁸ Finally, no transcriptional changes were detected for Vegfa, an angiogenesis factor released mainly by astrocytes, suggesting that the increase in astrocytic activity is not tied to increased angiogenesis at least at the molecular level.

Figure 2.

Transcriptional response and morphological changes following nucleus accumbens (NAc) infarct across time. A, Time-dependent expression of genes associated with endothelial cells (Pecam1; sham: day [D] 3: P=0.0027; D7: P≤0.0001; and D14: P=0.0005; ET-1 (endothelin-1): D3: P=0.5992; D7: P≤0.0001; and D14: P=0.0001), tight junctions (Cldn5: ET-1: D7: P=0.0005 and D14: P=0.0007; Ocln: ET-1: D14: P≤0.0001; Tjp1: sham: D3: P≤0.0001; D7: P=0.9822; and D14: P≤0.0001; and ET-1: D3: P≤0.0001; D7: P=0.6415; and D14: P≤0.0001), astrocyte reactivity (Gfap: ET-1: D7: P≤0.0001), and angiogenesis (Vegfa) in sham (left) or ET-1–injected mice (right). The dotted line represents normalized gene expression vs 1 day post-surgery for each gene. B, Gene expression of sham or ET-1–injected mice normalized to naive animals that did not undergo surgery (dotted line) for Pecam1, Cldn5, Ocln, Tjp1, Gfap, and Vegfa blood-brain barrier–related genes with significant effects for sham: Pecam1 (2-way ANOVA: surgery effect: P≤0.0001), Gfap (2-way ANOVA: surgery effect: P=0.0080), Vegfa (2-way ANOVA: surgery effect: P=0.0238), ET-1: Pecam1 (2-way ANOVA: stroke effect: naive vs ET-1: P=0.0011; stroke effect: sham vs ET-1: P=0.0003), and Vegfa (stroke effect: sham vs ET-1: P=0.0274). C, Immunostaining of Cldn5 (claudin-5) with the CD31 (cluster of differentiation 31) endothelial cell marker in sham vs ET-1–injected mice. Protein levels were normalized on naive animals, and an increase in Cldn5/CD31 ratio was noted for the ET-1 group at 7 and 14 days poststroke (2-way ANOVA: stroke effect: P=0.0185; n=3–4/group). D, Triple immunostaining with the cell-specific markers NeuN (neuronal nuclei), Gfap (glial fibrillary acidic protein), and Iba-1 (ionized calcium-binding adapter molecule 1) for neurons, astrocytes, and microglia, respectively, revealed gliosis in the NAc of ET-1–injected mice. Data represent mean±SEM. Two-way ANOVA followed by the Tuckey multiple comparison test was applied with ****P≤0.0001, **P≤0.01, and *P≤0.05.

To assess the impact of surgery and injection of fluid (ET-1 or sterile water for sham) on the BBB in the NAc, we normalized gene expression for sham or ET-1–injected mice at each timepoint to naive animals, which did not undergo surgery (Figure 2B). Unsurprisingly, a significant time effect was detected for all genes analyzed, indicating that the neurovascular gene expression is modulated throughout time after surgery. Significant surgery effects were observed for Pecam1, Gfap, and Vegfa with ET-1–mediated effects noted for Pecam1 and Vegfa. It suggests that injection of this vasoconstrictor in the male NAc affects mainly endothelial cells and angiogenesis. No treatment effect was observed for Cldn5, Ocln, and Tjp1 expressions. However, immunostaining revealed increased Cldn5/CD31 (cluster of differentiation 31) protein level poststroke in the NAc supporting healing processes (Figure 2C). In line with higher NAc Gfap level (Figure 2B), gliosis occurs following ET-1 injection with activation of astrocytes around the infarct and recruitment of microglia within the lesion (Figure 2D). Overall, our findings shed light on BBB-related distinct molecular and morphological profiles for surgery alone versus ET-1 injection, at least up to 14 days poststroke, and the implication of different cell types in insult recovery.

An Infarct in the NAc Impairs Sociability but Not Other Anxiety- or Depressive-Like Behaviors

To evaluate whether unilateral NAc infarct is sufficient to alter behaviors linked to anxiety and mood disorders, a battery of behavioral tests was conducted⁴⁹ on sham or ET-1–injected animals and performances compared with naive mice that were not subjected to surgery (Figure 3A). The mice were allowed to recover for 7 days poststroke because we observed most transcriptional changes at this timepoint (Figure 2). Naive, sham, or ET-1 injected mice were weighed daily and exposed to the elevated plus maze (anxiety), social interaction (SI) test (sociability), tail suspension test (depression), forced swim test (depression), or sucrose preference test (anhedonia, core MDD symptom; Figure 3A and 3B). While, as expected, naive animals steadily gained weight, mice from both sham and ET-1 groups lost weight until 4 days post-surgery (Figure 3B). No difference was observed between groups in the elevated plus maze for time spent in the maze with open arms, suggesting that an NAc infarct does not induce anxiety-like behaviors (Figure 3C, left). The total distance traveled was similar indicative of intact locomotor function (Figure 3C, right). To assess social interactions, mice were first allowed to freely explore an open field arena, and then, they were exposed to a novel social target during the second trial. This is a behavioral assessment protocol commonly used to evaluate the impact of stress exposure.^32,45 The SI ratio is calculated by dividing the time spent in the interaction zone when the social target is present divided by absent. Social interactions were significantly lower in both sham and ET-1–injected mice compared with naive mice, which was not due to locomotion deficits (Figure 3D). However, only 10% of sham mice display social avoidance with an SI ratio <1, while 42% of ET-1–injected mice preferred to avoid the social target, suggesting a negative effect of stroke in the NAc on social interactions. Surprisingly, both sham and ET-1 groups spent less time immobile in the tail suspension test versus naive mice (Figure 3E). This effect of surgery was not observed in the forced swim test, or sucrose preference test, also linked to a depressive phenotype (Figure 3F). Altogether, these results suggest that ET-1–induced stroke in the NAc of male mice impairs sociability but not other behavioral domains associated with anxiety or depression.

Figure 3.

Assessment of social, anxiety, and depressive-like behaviors in male mice following ET-1 (endothelin-1)–induced unilateral infarct in the nucleus accumbens (NAc). A, Experimental timeline and groups. B, Changes in body weight across time for naive (light gray), sham (dark gray), and ET-1 (red) mice (n=11–12/group). C, Time spent in the open arms of the elevated plus maze, distance traveled, and representative heatmaps (n=11–12/group). D, Social interaction (SI) ratios (1-way ANOVA: treatment effect: P=0.0006; sham: P=0.0281; and ET-1: P=0.0039), distance traveled, and pie charts representing proportion of mice considered avoidant (SI ratio <1, dark blue) or social (SI ratio >1, light blue) for each group along with representative heatmap of time spent in the arena during the second trial (n=11–12/group). E, Time spent immobile in the tail suspension test (1-way ANOVA: treatment effect: P=0.0021; sham: P=0.0062; and ET: P=0.0053; n=11–12/group). F, Time spent immobile in the forced swim test (left) and sucrose preference over 48 hours (right; n=11–12/group). Data represent mean±SEM. One-way ANOVA followed by the Tuckey multiple comparison test was applied with ****P≤0.0001, ***P≤0.001, **P≤0.01, and *P≤0.05.

Discussion

One-third of individuals experiencing a stroke will experience PSD leading to poorer neurological outcomes⁵⁰ and increased hospitalization costs.⁵¹ Although strokes restricted to the NAc are rare, we targeted this region due to its important role in reward, mood, and emotional processing. Focal ischemic lesions are difficult to achieve due to the localization of the region of interest. Happ et al²¹ previously attempted to characterize ET-1–induced infarct in the NAc in rats but failed due to a high mortality rate. ET-1 injection in the cerebrospinal fluid was shown to cause potential respiratory insufficiency.⁵² Here, optimization of Bregma coordinates was necessary to avoid infiltration of ET-1 in the ventricles, resulting in high survival rates. Arteries lateral to the NAc also had to be avoided to prevent nonfocal lesions.^53,54 A limitation is that only ≈50% of the lesion was in the NAc with needle track injury contributing at least for early timepoints.

Ischemic lesions induce vascular remodeling in the early stages of recovery with BBB opening lasting up to 72 hours.⁵⁵ We showed that viral-mediated targeted BBB opening is sufficient to induce a depressive-like phenotype.³² Because BBB disruption is associated with both stroke and MDD, it could play a central role in PSD biology. We assessed transcriptional changes in BBB-associated genes up to 14 days following the ischemic lesion, revealing that sham and ET-1–injected mice display both similar and distinct neurovascular responses. In MCAO models, increased vascularization is noted up to 1 week following the ischemic lesion.^56,57 We did not observe a change in the expression of Vegfa, a vascular growth factor essential for angiogenesis, suggesting a discrepancy with MCAO studies.⁵⁸

Tight junctions have been shown to be impacted in a biphasic manner shortly following stroke.³⁴ In our case, tight junction expression is modulated even after a sham surgery and, thus, not by the lesion itself. This could be due to the timepoint chosen for analysis. Nevertheless, our transcriptomic results suggest that BBB-associated genes can be regulated by surgery with ET-1 injection inducing a distinct pattern of molecular changes mostly in endothelial cells. Moreover, we noted increased Cldn5/CD31 protein levels in ET-1–injected mice at 7 and 14 days poststroke compared with sham animals supporting healing processes. Isoflurane can disrupt BBB properties.^59,60 It was the anesthetic of choice for our study, and we cannot rule out that it had long-lasting effects on tight junction expression. Future studies should be conducted to evaluate in detail and at multiple timepoints morphological adaptations after ET-1 injection in the NAc.

The astrocytic marker, Gfap, was also modulated by surgery alone. Astrocyte activation following brain lesions has been reported before in various contexts including stroke.^48,61 Here, NAc ET-1 lesion seems to promote stronger reactivity of these glial cells and microglia recruitment in this brain area compared with the sham injection, such as in the MCAO model.⁵⁶ Brain inflammation has been linked to the establishment of social avoidance and depression-like behaviors with neuroimmune mechanisms of depression receiving increasing attention.^62,63 Deciphering the contribution of the different glial and BBB cells in PSD may help identify novel targets to design innovative therapies to treat this condition.

At the behavioral level, a small and transient unilateral lesion in the NAc was not sufficient to induce a PSD phenotype. Nevertheless, it altered sociability, a critical feature of human life and well-being. Only a few models of stroke can promote depressive- and anxiety-like behaviors by themselves.⁶⁴ Vahid-Ansari et al⁴⁰ successfully induced behavioral deficits 1 and 6 weeks post-infarct after ET-1 injection in the prefrontal cortex of mice, a hub for cognitive processes, emotion regulation, and decision-making. Kronenberg et al⁶⁵ reported behavioral changes after MCAO, prevented by treatment with the antidepressant citalopram. Most PSD models combine an ischemic lesion with a chronic stressor,⁶⁴ commonly used to mimic MDD in rodents.⁶⁶ It would be relevant to perform a stress paradigm following an NAc infarct in future studies and evaluate how it affects BBB properties along with neuronal function. Intriguingly, both MDD and PSD are more prevalent in women,^13,67 and sex differences in BBB dysfunction have been reported in MDD.^32,41,68 It will be important to investigate potential sex differences in the vascular pathophysiology of PSD.

To sum up, our study highlights the technical complexity of performing focal lesions in a deep brain area such as the NAc. Despite the fact that NAc ET-1 injection did not induce a full PSD phenotype, it affected sociability even without stress exposure, which is promising to better understand the biology underlying the apparition of depressive symptoms after stroke. Increasing evidence supports an involvement of neurovascular changes in MDD,⁶⁹ and with BBB alterations consistently observed in stroke, it is important to decipher if and how vascular-related cells are engaged in PSD pathophysiology.

Article Information

Acknowledgments

The authors thank Céline Leclerc, Daphnée Le Sage, and Marie-Laurence Turmel for their technical help and expertise essential to collect the magnetic resonance imaging scans and the CERVO Brain Research Center Housing Facility Staff (special thanks to Louisabelle Gagnon) for their work and support.

Sources of Funding

This research was supported by the Canadian Institutes for Health Research (CIHR; project grant 427011 to Dr Ménard), the Canada First Research Excellence Fund (Sentinel North Research Chair to Dr Ménard), and Fonds de Recherche du Québec-Santé (FRQS; Junior 2 Salary Award to Dr Ménard). Drs Bouchard, A. Collignon, L.B. Binder, L.M. Bevilacqua, and Dr Dion-Albert are supported by MSc and PhD scholarships from CIHR, FRQS, and NeuroQuébec.

Disclosures

None.

Supplemental Material

Supplemental Methods

Table S1

Figure S1

ARRIVE Checklist