Transfer of mitochondria from astrocytes to neurons after stroke.

Neurons can release damaged mitochondria and transfer them to astrocytes for disposal and recycling. This ability to exchange mitochondria may represent a potential mode of cell-to-cell signalling in the central nervous system. Here we show that astrocytes in mice can also release functional mitochondria that enter neurons. Astrocytic release of extracellular mitochondrial particles was mediated by a calcium-dependent mechanism involving CD38 and cyclic ADP ribose signalling. Transient focal cerebral ischaemia in mice induced entry of astrocytic mitochondria into adjacent neurons, and this entry amplified cell survival signals. Suppression of CD38 signalling by short interfering RNA reduced extracellular mitochondria transfer and worsened neurological outcomes. These findings suggest a new mitochondrial mechanism of neuroglial crosstalk that may contribute to endogenous neuroprotective and neurorecovery mechanisms after stroke.

Recurrent Stroke Due to Metastatic Pulmonary Tumor Emboli as an Important Clinical Entity.

We present an autopsy case of repetitive stroke due to tumor emboli, indistinguishable from thromboembolism with a hypercoagulable state in its clinical course. A 72-year-old man diagnosed with stage IVA oropharyngeal squamous cell carcinoma received chemoradiotherapy. Follow-up imaging revealed mediastinal lymph nodes and pulmonary metastasis. One year later, the patient experienced right arm weakness, and brain magnetic resonance imaging showed acute ischemic lesions in multiple vascular territories. He was diagnosed with paradoxical cerebral embolism due to cancer-associated venous thrombosis and treated with rivaroxaban. However, newly developed cerebral infarcts were confirmed 1 month later. Then, rivaroxaban treatment was switched to subcutaneous unfractionated heparin injection. He was admitted again for stroke recurrence and died of respiratory failure 8 days after admission. Autopsy demonstrated pulmonary metastasis invading the veins and tumor emboli in the culprit cerebral arteries. D-dimer was kept constant at a slightly higher level, ranging from 1 to 3 µg/mL during the course of recurrence. We should consider tumor embolism in the differential diagnosis of recurrent stroke along with pulmonary tumor and resistance to heparin preparations with unchanged D-dimer levels.

Delayed inhibition of c-Jun N-terminal kinase worsens outcomes after focal cerebral ischemia.

The stress-activated protein kinase c-Jun N-terminal kinase (JNK) is a central regulator in neuronal death cascades. In animal models of cerebral ischemia, acute inhibition of JNK reduces infarction and improves outcomes. Recently however, emerging data suggest that many neuronal death mediators may have biphasic properties-deleterious in the acute stage but potentially beneficial in the delayed stage. Here, we hypothesized that JNK may also have biphasic actions, so some caution may be required in the development of JNK inhibitors for stroke. Sprague Dawley rats underwent 90 min transient occlusions of the middle cerebral artery. Acute treatment (10 min poststroke) with the JNK inhibitor SP600125 reduced infarction volumes. In contrast, delayed treatment (7 d poststroke) worsened infarction volumes and neurological outcomes. Immunostaining of peri-infarct cortex showed that JNK inhibition suppressed surrogate markers of neurovascular remodeling, including matrix metalloproteinase-9 in GFAP-positive astrocytes and microvascular density. Consistent with these in vivo data, SP600125 significantly suppressed in vitro angiogenesis in rat brain endothelial cultures. Our data provide initial proof-of-concept that the neuronal death target JNK may also participate in endogenous processes of neurovascular remodeling and recovery after cerebral ischemia.

Granulocyte colony-stimulating factor enhances arteriogenesis and ameliorates cerebral damage in a mouse model of ischemic stroke.

BACKGROUND AND PURPOSE: Enhancing collateral artery growth is a potent therapeutic approach to treat cardiovascular ischemic disease from occlusive artery. Granulocyte-macrophage colony-stimulating factor (GM-CSF) has gained attention for its ability to promote arteriogenesis, ameliorating brain damage, by the mechanisms involving monocyte upregulation. However, the recent clinical study testing its efficacy in myocardial ischemia has raised the question about its safety. We tested alternative colony-stimulating factors for their effects on collateral artery growth and brain protection. METHODS: Brain hypoperfusion was produced by occluding the left common carotid artery in C57/BL6 mice. After the surgery, granulocyte colony-stimulating factor, macrophage colony-stimulating factor, or GM-CSF (100 µg/kg/day) was administered daily for 5 days. The angioarchitecture for leptomeningeal anastomoses and the circle of Willis were visualized after the colony-stimulating factor treatment. Circulating blood monocytes and Mac-2-positive cells in the dorsal surface of the brain were determined. A set of animals underwent subsequent ipsilateral middle cerebral artery occlusion and infarct volume was assessed. RESULTS: Granulocyte colony-stimulating factor as well as GM-CSF promoted leptomeningeal collateral growth after common carotid artery occlusion. Both granulocyte colony-stimulating factor and GM-CSF increased circulating blood monocytes and Mac-2-positive cells in the dorsal brain surface, suggesting the mechanisms coupled to monocyte upregulation might be shared. Infarct volume after middle cerebral artery occlusion was reduced by granulocyte colony-stimulating factor, similarly to GM-CSF. Macrophage colony-stimulating factor showed none of theses effects. CONCLUSIONS: Granulocyte colony-stimulating factor enhances collateral artery growth and reduces infarct volume in a mouse model of brain ischemia, similarly to GM-CSF.

Cilostazol, not aspirin, reduces ischemic brain injury via endothelial protection in spontaneously hypertensive rats.

BACKGROUND AND PURPOSE: It is well-established that hypertension leads to endothelial dysfunction in the cerebral artery. Recently, cilostazol has been used for the secondary prevention of ischemic stroke. Among antiplatelet drugs, phosphodiesterase inhibitors including cilostazol have been shown to have protective effects on endothelial cells. The aim of the present study is to investigate the effects of cilostazol and aspirin on endothelial nitric oxide synthase (eNOS) phosphorylation in the cerebral cortex, endothelial function, and infarct size after brain ischemia in spontaneously hypertensive rats (SHR). METHODS: Five-week-old male SHR received a 5-week regimen of chow containing 0.1% aspirin, 0.1% cilostazol, 0.3% cilostazol, or the vehicle control. The levels of total and Ser(1177)-phosphorylated eNOS protein in the cerebral cortex were evaluated by Western blot. To assess the contribution of eNOS in maintaining cerebral blood flow, we monitored cerebral blood flow by laser-Doppler flowmetry after L-N(5)-(1-iminoethyl)ornithine infusion. Additionally, we evaluated residual microperfusion using fluorescence-labeled serum protein and infarct size after transient focal brain ischemia. RESULTS: In SHR, the blood pressure and heart rate were similar among the groups. Cilostazol-treated SHR had a significantly higher ratio of phospho-eNOS/total eNOS protein than vehicle-treated and aspirin-treated SHR. Treating with cilostazol, but not aspirin, significantly improved cerebral blood flow response to L-N(5)-(1-iminoethyl)ornithine. Cilostazol also increased residual perfusion of the microcirculation and reduced brain damage after ischemia compared to vehicle control and aspirin. CONCLUSIONS: These findings indicate that cilostazol, but not aspirin, can attenuate ischemic brain injury by maintaining endothelial function in the cerebral cortex of SHR.

Hypertension impairs leptomeningeal collateral growth after common carotid artery occlusion: restoration by antihypertensive treatment.

Chronic mild hypoperfusion has been shown to enlarge pial collateral vessels in normal mouse brains. The purpose of this study was to clarify the effect of hypertension on pial collateral vessel development after chronic hypoperfusion using spontaneously hypertensive rats (SHR). In normotensive rats, unilateral common carotid artery (CCA) occlusion enlarged leptomeningeal collateral vessels. CCA occlusion also preserved residual cerebral blood flow (CBF) and attenuated infarct size after middle cerebral artery (MCA) occlusion 14 days later. In contrast, in SHR, CCA occlusion neither enlarged the leptomeningeal anastomosis nor showed protective effects after MCA occlusion. However, decreasing blood pressure using an angiotensin II AT1 receptor blocker restored the beneficial effect of CCA occlusion on collateral growth as well as on residual CBF and infarct size after MCA occlusion. Adaptive responses in CBF autoregulation curves observed 14 days after CCA occlusion in normotensive rats were impaired in untreated SHR, but were restored after antihypertensive treatment. In conclusion, SHR have impaired leptomeningeal collateral growth after CCA occlusion, but antihypertensive treatment restores the beneficial effect of CCA occlusion on collateral circulation.

An angiotensin II type 1 receptor blocker can preserve endothelial function and attenuate brain ischemic damage in spontaneously hypertensive rats.

Hypertension reduces endothelial nitric oxide synthase (eNOS) expression and leads to endothelial dysfunction. However, few studies have demonstrated the influences of hypertension on eNOS function in the cerebral cortex. The present study investigates the influences of hypertension on endothelial function in the cerebral cortex and the protective effects of antihypertensive agents against brain ischemia through the preservation of endothelial function. Five- and ten-week-old male Wistar rats and spontaneously hypertensive rats (SHR) were used for experiments. Five-week-old SHR received olmesartan, hydralazine, or vehicle for 5 weeks in drinking water. eNOS activation in the cerebral cortex was evaluated by analyzing levels of total and Ser(1177)-phosphorylated eNOS protein by Western blot. Blood pressure of 10-week-old SHR without treatment was clearly high, and the ratio of phospho-eNOS/total eNOS protein was significantly low. Five-week treatment with olmesartan or hydralazine suppressed the elevation of blood pressure and the reduction of phosphorylated eNOS-Ser(1177) in SHR, and olmesartan was more effective in maintaining phosphorylation of eNOS-Ser(1177) than hydralazine. To assess the contribution of eNOS to maintaining cerebral blood flow (CBF), we monitored CBF by laser-Doppler flowmetry after L-N(5)-(1-iminoethyl)ornithine (L-NIO) infusion. CBF response to L-NIO was preserved in olmesartan-treated SHR but not in hydralazine-treated SHR. Furthermore, infarct volume 48 hr after transient focal brain ischemia in olmesartan-treated SHR was significantly reduced compared with vehicle-treated SHR. These findings indicate that chronic prehypertensive treatment with olmesartan could attenuate brain ischemic injury through the maintenance of endothelial function in the cerebral cortex in SHR.

Clinical significance of oral intake in patients with acute stroke.

We investigated the feeding method and predictors for oral intake difficulty for a month after acute stroke. In 107 consecutive patients, swallowing function was assessed using a bedside screening protocol within 48 h of admission. The method of feeding was followed for 4 weeks, and predictors for "non-oral intake" on admission and 4 weeks later were analyzed. Sixty-two patients (58%) were fed any type of food orally within 48 h of admission, and 91 patients (84%) were fed orally 4 weeks later. Independent predictors for non-oral intake within 48 h of admission were consciousness disturbance (not completely alert; OR = 12.3), absence of gag reflex (OR = 5.34), and NIHSS score (OR = 1.20 per one point). Independent predictors for non-oral intake after 4 weeks were absence of gag reflex (OR = 7.95) and NIHSS score (OR = 1.13 per one point) on admission. Only four (9%) patients in the non-oral intake group within 48 h of admission and no patients in the non-oral intake group 4 weeks after admission were discharged to home. In acute stroke patients, absence of the gag reflex and severe neurologic deficits on admission predict prolonged dysphagia lasting longer than a month. Patients who could not eat orally had poor outcome.

Transcriptomic characterization of microglia activation in a rat model of ischemic stroke.

Microglia are key regulators of inflammatory response after stroke and brain injury. To better understand activation of microglia as well as their phenotypic diversity after ischemic stroke, we profiled the transcriptome of microglia after 75 min transient focal cerebral ischemia in 3-month- and 12-month-old male spontaneously hypertensive rats. Microglia were isolated from the brains by FACS sorting on days 3 and 14 after cerebral ischemia. GeneChip Rat 1.0ST microarray was used to profile the whole transcriptome of sorted microglia. We identified an evolving and complex pattern of activation from 3 to 14 days after stroke onset. M2-like patterns were extensively and persistently upregulated over time. M1-like patterns were only mildly upregulated, mostly at day 14. Younger 3-month-old brains showed a larger microglial response in both pro- and anti-inflammatory pathways, compared to older 12-month-old brains. Importantly, our data revealed that after stroke, most microglia are activated towards a wide spectrum of novel polarization states beyond the standard M1/M2 dichotomy, especially in pathways related to TLR2 and dietary fatty acid signaling. Finally, classes of transcription factors that might potentially regulate microglial activation were identified. These findings should provide a comprehensive database for dissecting microglial mechanisms and pursuing neuroinflammation targets for acute ischemic stroke.

Granulocyte-macrophage colony-stimulating factor enhances leptomeningeal collateral growth induced by common carotid artery occlusion.

BACKGROUND AND PURPOSE: Granulocyte-macrophage colony-stimulating factor (GM-CSF) has been reported to accelerate collateral growth (arteriogenesis) at the circle of Willis in rat brain. However, the effect of GM-CSF on leptomeningeal collateral growth has not been established. We examined the effect of unilateral common carotid artery (CCA) occlusion and GM-CSF treatment on leptomeningeal collateral growth in mice. METHODS: Adult mice were subjected to unilateral CCA occlusion or sham surgery followed by an alternate-day regimen of GM-CSF (20 microg/kg) or saline injection. On day 7, latex perfusion was performed in 1 set of mice to visualize the leptomeningeal vessels, and the number of Mac-2(+) monocytes/macrophages on the dorsal surface of the brain was counted. In another set of mice, on day 7, permanent ipsilateral middle cerebral artery (MCA) occlusion was performed, and infarct volume was measured. RESULTS: Leptomeningeal collateral growth was observed after CCA occlusion, and that was enhanced by GM-CSF treatment. An increase in the number of Mac-2(+) cells on the surface of the brain occurred after CCA occlusion and was enhanced by GM-CSF treatment. Seven days after CCA occlusion, GM-CSF treatment decreased the infarct size attributable to subsequent MCA occlusion. CONCLUSIONS: After CCA occlusion, GM-CSF treatment enhanced leptomeningeal collateral growth and decreased the infarct size after MCA occlusion in mice.

The phosphodiesterase inhibitor rolipram promotes survival of newborn hippocampal neurons after ischemia.

BACKGROUND AND PURPOSE: Brain ischemia stimulates neurogenesis. However, newborn neurons show a progressive decrease in number over time. Under normal conditions, the cAMP-cAMP responsive element binding protein (CREB) pathway regulates the survival of newborn neurons. Constitutive activation of CREB after brain ischemia also stimulates hippocampal neurogenesis. Thus, activation of cAMP-CREB signaling may provide a promising strategy for enhancing the survival of newborn neurons. We examined whether treatment of mice with the phosphodiesterase-4 inhibitor rolipram enhances hippocampal neurogenesis after ischemia. METHODS: Both common carotid arteries in mice were occluded for 12 minutes. Bromodeoxyuridine (BrdU) was used to label proliferating cells. Mice were perfused transcardially with 4% paraformaldehyde, and immunohistochemistry was performed. To evaluate the role of CREB in the survival of newborn neurons after ischemia, intrahippocampal injection of a CRE-decoy oligonucleotide was delivered for 1 week. We examined whether the activation of cAMP-CREB signaling by rolipram enhanced the proliferation and survival of newborn neurons. RESULTS: Phospho-CREB immunostaining was markedly upregulated in immature neurons, decreasing to low levels in mature neurons. The number of BrdU-positive cells 30 days after ischemia was significantly less in the CRE-decoy treatment group than in the vehicle group. Rolipram enhanced the proliferation of newborn cells under physiologic conditions but not under ischemic conditions. Rolipram significantly increased the survival of nascent BrdU-positive neurons, accompanied by an enhancement of phospho-CREB staining and decreased newborn cell death after ischemia. CONCLUSIONS: CREB phosphorylation regulates the survival of newborn neurons after ischemia. Chronic pharmacological activation of cAMP-CREB signaling may be therapeutically useful for the enhancement of neurogenesis after ischemia.

Postischemic administration of HMG CoA reductase inhibitor inhibits infarct expansion after transient middle cerebral artery occlusion.

Infarcts expand over a period of several days after focal cerebral ischemia. Inhibition of infarct expansion would be a promising strategy for the protection of the brain after ischemic insult. In this study, we examined the effect of statin treatment initiated 2 days after transient occlusion of the middle cerebral artery for 80 min. Treatment with continuous infusion of pravastatin with an osmotic minipump for 6 days resulted in a significantly smaller infarct volume and better recovery of motor function at 8 days after ischemia compared to vehicle treatment. These results suggest that statin treatment during the postischemic period may be effective in stroke patients.

Postischemic exercise decreases neurogenesis in the adult rat dentate gyrus.

Running exercise enhances neurogenesis in the normal adult and aged hippocampus. However, the effect of exercise on neurogenesis in the ischemic hippocampus is unclear. Here, we show that running exercise has different effects on ischemic and non-ischemic brain. Young (3-4-month-old) normotensive Wistar rats were used for this study. We administered bromodeoxyuridine (BrdU) to rats 7 days after the induction of transient forebrain ischemia or sham operation. BrdU-labeled cells were increased in the ischemic subgranular zone (SGZ) and granule cell layer (GCL) and double immunofluoresence showed approximately 80% of BrdU-labeled cells expressed neuronal markers. To assess the effect of running exercise on neurogenesis, BrdU-labeled cells in these regions were quantified after 1 day and 14 days. In sham-operated rats, the numbers of BrdU-labeled cells were significantly increased (2.2-fold) in the SGZ and GCL in response to running exercise. The numbers of BrdU-labeled cells were increased in response to ischemia, however, they were decreased 14 days after BrdU administration and running exercise accelerated the reduction in BrdU-labeled cells in ischemic rats. These findings suggest that running exercise has a negative effect on neurogenesis in the ischemic hippocampus. This may be important with respect to assessment of therapeutic approaches for functional recovery after stroke.

Characteristics of cryptogenic stroke in cancer patients.

OBJECTIVE: To clarify the characteristics of cryptogenic stroke in patients with active cancer. METHODS: Patients with or without cancer diagnosed with acute ischemic stroke between January 2006 and February 2015 were extracted from a prospectively collected stroke database of Osaka University Hospital. Patients were categorized according to the presence of active cancer and known stroke mechanisms. RESULTS: Among 1191 patients with acute ischemic stroke, 145 (12%) had active cancer. Patients with active cancer were diagnosed more often with cryptogenic stroke than were patients without cancer (47% vs. 12%, P < 0.001). Compared with cryptogenic stroke patients without cancer, cryptogenic stroke patients with active cancer had fewer atherosclerotic risk factors, lower nutrition status, higher plasma D-dimer levels, and multiple vascular lesions. In a multivariate logistic analysis, plasma D-dimer level (odds ratio [OR] per 1 standard deviation increase: 6.30; 95% confidence interval [CI]: 2.94-15.69; P < 0.001), and the presence of multiple vascular lesions (OR: 6.40; 95% CI: 2.35-18.35; P < 0.001) were independent predictors of active cancer. When comparing active cancer patients who had known stroke mechanisms with those who had cryptogenic stroke, high plasma D-dimer levels, multiple vascular lesions, and receiving chemotherapy and/or radiation therapy were associated with cryptogenic stroke etiology. INTERPRETATION: In cryptogenic stroke, patients with active cancer has a unique pathology characterized by high plasma D-dimer levels and multiple vascular lesions. The hypercoagulable state and malnutrition due to cancer and its treatments potentially influence the development of cryptogenic stroke in cancer patients.

CD200 restrains macrophage attack on oligodendrocyte precursors via toll-like receptor 4 downregulation.

There are numerous barriers to white matter repair after central nervous system injury and the underlying mechanisms remain to be fully understood. In this study, we propose the hypothesis that inflammatory macrophages in damaged white matter attack oligodendrocyte precursor cells via toll-like receptor 4 signaling thus interfering with this endogenous progenitor recovery mechanism. Primary cell culture experiments demonstrate that peritoneal macrophages can attack and digest oligodendrocyte precursor cells via toll-like receptor 4 signaling, and this phagocytosis of oligodendrocyte precursor cells can be inhibited by using CD200-Fc to downregulate toll-like receptor 4. In an in vivo model of white matter ischemia induced by endothelin-1, treatment with CD200-Fc suppressed toll-like receptor 4 expression in peripherally circulating macrophages, thus restraining macrophage phagocytosis of oligodendrocyte precursor cells and leading to improved myelination. Taken together, these findings suggest that deleterious macrophage effects may occur after white matter ischemia, whereby macrophages attack oligodendrocyte precursor cells and interfere with endogenous recovery responses. Targeting this pathway with CD200 may offer a novel therapeutic approach to amplify endogenous oligodendrocyte precursor cell-mediated repair of white matter damage in mammalian brain.

[A case of rheumatoid meningitis presented with generalized seizure in whom MRI images were helpful for the diagnosis].

We report a 65-years-old woman with rheumatoid meningitis presented with a generalized seizure. She has a 18-year history of rheumatoid arthritis, which has been successfully treated. She developed a generalized seizure. She was diagnosed as having subarachnoid hemorrhage, because the brain magnetic resonance imaging (MRI) showed increased fluid attenuated inversion recovery (FLAIR) signals in her left frontoparietal subarachnoid space. After one month of clinical stabilization, she developed numbness and weakness in her right lower extremity that spread to her right upper extremity and face. Brain MRI showed progression of subarachnoid lesion on FLAIR image and leptomeningeal enhancement on gadolinium-enhanced T1 weighted image. She was diagnosed as having rheumatoid meningitis, and methylprednisolone pulse therapy was started. Then, her symptoms and MRI findings were rapidly improved. Though rheumatoid meningitis is rare and presents a difficulty in the diagnosis, MRI features may support the diagnosis.

Three-Dimensional Blood-Brain Barrier Model for in vitro Studies of Neurovascular Pathology.

Blood-brain barrier (BBB) pathology leads to neurovascular disorders and is an important target for therapies. However, the study of BBB pathology is difficult in the absence of models that are simple and relevant. In vivo animal models are highly relevant, however they are hampered by complex, multi-cellular interactions that are difficult to decouple. In vitro models of BBB are simpler, however they have limited functionality and relevance to disease processes. To address these limitations, we developed a 3-dimensional (3D) model of BBB on a microfluidic platform. We verified the tightness of the BBB by showing its ability to reduce the leakage of dyes and to block the transmigration of immune cells towards chemoattractants. Moreover, we verified the localization at endothelial cell boundaries of ZO-1 and VE-Cadherin, two components of tight and adherens junctions. To validate the functionality of the BBB model, we probed its disruption by neuro-inflammation mediators and ischemic conditions and measured the protective function of antioxidant and ROCK-inhibitor treatments. Overall, our 3D BBB model provides a robust platform, adequate for detailed functional studies of BBB and for the screening of BBB-targeting drugs in neurological diseases.

Potential interactions between pericytes and oligodendrocyte precursor cells in perivascular regions of cerebral white matter.

Pericytes are embedded within basal lamina and play multiple roles in the perivascular niche in brain. Recently, oligodendrocyte precursor cells (OPCs) have also been reported to associate with cerebral endothelium. Is it possible that within this gliovascular locus, there may also exist potential spatial and functional interactions between pericytes and OPCs? Here, we demonstrated that in the perivascular region of cerebral white matter, pericytes and OPCs may attach and support each other. Immunostaining showed that pericytes and OPCs are localized in close contact with each other in mouse white matter at postnatal days 0, 60 and 240. Electron microscopic analysis confirmed that pericytes attached to OPCs via basal lamina in the perivascular region. The close proximity between these two cell types was also observed in postmortem human brains. Functional interaction between pericytes and OPCs was assessed by in vitro media transfer experiments. When OPC cultures were treated with pericyte-conditioned media, OPC number increased. Similarly, pericyte number increased when pericytes were maintained in OPC-conditioned media. Taken together, our data suggest a potential anatomical and functional interaction between pericytes and OPCs in cerebral white matter.

Granulocyte colony-stimulating factor fails to enhance leptomeningeal collateral growth in spontaneously hypertensive rats.

The promotion of collateral artery growth is an attractive approach for the treatment of chronic brain hypoperfusion due to occlusive artery disease. We previously reported that hypertension impaired the collateral artery growth of leptomeningeal anastomoses after brain hypoperfusion. Granulocyte colony-stimulating factor (G-CSF) enhances arteriogenesis in a mouse model via a mechanism involving monocyte/macrophage mobilization. However, the arteriogenic effect of G-CSF in hypertension remains unknown. In the present study, we tested whether G-CSF affected collateral artery growth in both normotensive and hypertensive model rat. Left common carotid artery (CCA) occlusion was performed to induce hypoperfusion in the brains of Wistar rats and spontaneously hypertensive rats (SHR). G-CSF was administered subcutaneously for 5 consecutive days. The superficial angioarchitecture of the leptomeningeal anastomoses and the circle of Willis after CCA occlusion and G-CSF treatment were visualized by latex perfusion. Circulating blood monocytes and CD68-positive cells, which represented the macrophages on the dorsal surface of the brain, were counted. G-CSF enhanced leptomeningeal collateral growth in Wistar rats, but not in SHR. G-CSF increased circulating blood monocytes in both Wistar rats and SHR. The number of CD68-positive cells on the dorsal surface of the brain was increased by G-CSF in Wistar rats, but not in SHR. The increase in macrophage accumulation correlated with the observed arteriogenic effects. In conclusion, G-CSF promotes collateral artery growth in the normotensive model rat, but not in the hypertensive model rat.