Blood flow into the brain is dynamically regulated to satisfy the changing metabolic requirements of neurons, but how this is accomplished has remained unclear. Here, we demonstrate a central role for capillary endothelial cells in sensing neural activity and communicating it to upstream arterioles in the form of an electrical vasodilatory signal. We further demonstrate that this signal is initiated by extracellular potassium (K+)—a byproduct of neural activity—which activates capillary endothelial cell inward-rectifier K+ (KIR2.1) channels to produce a rapidly propagating retrograde hyperpolarization that causes upstream arteriolar dilation, increasing blood flow into the capillary bed. Our results establish brain capillaries as an active sensory web that converts changes in external K+ into rapid, ‘inside-out’ electrical signaling to direct blood flow to active brain regions.

The cellular events that cause ischemic neurological damage following aneurysmal subarachnoid hemorrhage (SAH) have remained elusive. We report that subarachnoid blood profoundly impacts communication within the neurovascular unit-neurons, astrocytes, and arterioles-causing inversion of neurovascular coupling. Elevation of astrocytic endfoot Ca 2+ to ∼400 nM by neuronal stimulation or to ∼300 nM by Ca 2+ uncaging dilated parenchymal arterioles in control brain slices but caused vasoconstriction in post-SAH brain slices. Inhibition of K + efflux via astrocytic endfoot large-conductance Ca 2+ -activated K + (BK) channels prevented both neurally evoked vasodilation (control) and vasoconstriction (SAH

Since neural stem cells (NSCs) have the ability to migrate toward a tumor mass, genetically engineered NSCs were used for the treatment of gliomas. We first evaluated the ''bystander effect'' between NSCs transduced with the herpes simplex virus-thymidine kinase (HSVtk) gene (NSCtk) and C6 rat glioma cells under both in vitro and in vivo conditions. A potent bystander effect was observed in co-culture experiments of NSCtk and C6 cells. In the intracranial co-implantation experiments in athymic nude mice and Sprague-Dawley rats, the animals co-implanted with NSCtk and C6 cells and treated with ganciclovir (GCV) showed no intracranial tumors and survived more than 100 days, while those treated with physiological saline (PS) died of tumor progression. We next injected NSCtk cells into the pre-existing C6 tumor in rats and treated them with GCV or PS. The tumor volume was serially measured by magnetic resonance imaging. The tumor disappeared in six out of nine rats in the NSCtk/GCV group, while all the rats treated with PS died of tumor progression by day 21. The results indicate the feasibility of a novel gene therapy strategy for gliomas through a bystander effect generated by intratumoral injection of NSCtk cells and systemic GCV administration.

Subarachnoid hemorrhage (SAH) following cerebral aneurysm rupture is associated with high rates of morbidity and mortality. Surviving SAH patients often suffer from neurological impairment, yet little is currently known regarding the influence of subarachnoid blood on brain parenchyma. The objective of the present study was to examine the impact of subarachnoid blood on glial cells using a rabbit SAH model. The astrocyte-specific proteins, glial fibrillary acidic protein (GFAP) and S100B, were up-regulated in brainstem from SAH model rabbits, consistent with the development of reactive astrogliosis. In addition to reactive astrogliosis, cytosolic expression of the pro-inflammatory cytokine, high-mobility group box 1 protein (HMGB1) was increased in brain from SAH animals. We found that greater than 90% of cells expressing cytosolic HMGB1 immunostained positively for Iba1, a specific marker for microglia and macrophages. Further, the number of Iba1-positive cells was similar in brain from control and SAH animals, suggesting the majority of these cells were likely resident microglial cells rather than infiltrating macrophages. These observations demonstrate SAH impacts brain parenchyma by

Migraine is a debilitating neurological disorder characterized by mild to severe headache that is often accompanied by aura and other neurological symptoms. Among proposed mechanisms, dilation of the dural vasculature especially the middle meningeal artery (MMA) has been implicated as one component underlying this disorder. Several regulatory peptides from trigeminal sensory and sphenopalatine postganglionic parasympathetic fibers innervating these vessels have been implicated in the process including pituitary adenylate cyclase-activating polypeptide (PACAP). Although PACAP has been well described as a potent dilator in many vascular beds, the effects of PACAP on the dural vasculature are unclear. In the current study, we examined the ability of PACAP to dilate MMAs that were isolated from rats and pressurized ex vivo. PACAP38 potently dilated pressurized MMAs with an EC50 of 1 pM. The PAC1 receptor antagonist, PACAP(6-38), abolished MMA dilation caused by picomolar concentrations of PACAP. In contrast, cerebellar arteries isolated from the brain surface were ~1,000-fold less sensitive to PACAP than MMAs. Although cerebellar arteries expressed transcripts for all three PACAP receptor subtypes (PAC1, VPAC1, and VPAC2 receptors) by RT-PCR analyses, MMA demonstrated only PAC1 and VPAC2 receptor expression. Further, multiple variants of the PAC1 receptor were identified in the MMA. The expression of PAC1 receptors and the high potency of PACAP to induce MMA vasodilation are consistent with their potential roles in the etiology of migraine.

Subarachnoid hemorrhage (SAH) induces acute changes in the cerebral microcirculation. Recent findings ex vivo suggest neurovascular coupling (NVC), the process that increases cerebral blood flow upon neuronal activity, is also impaired after SAH. The aim of the current study was to investigate whether this occurs also in vivo. C57BL/6 mice were subjected to either sham surgery or SAH by filament perforation. Twenty-four hours later NVC was tested by forepaw stimulation and CO reactivity by inhalation of 10% CO. Vessel diameter was assessed in vivo by two-photon microscopy. NVC was also investigated ex vivo using brain slices. Cerebral arterioles of sham-operated mice dilated to 130% of baseline upon CO inhalation or forepaw stimulation and cerebral blood flow (CBF) increased. Following SAH, however, CO reactivity was completely lost and the majority of cerebral arterioles showed paradoxical constriction in vivo and ex vivo resulting in a reduced CBF response. As previous results showed intact NVC 3 h after SAH, the current findings indicate that impairment of NVC after cerebral hemorrhage occurs secondarily and is progressive. Since neuronal activity-induced vasoconstriction (inverse NVC) is likely to further aggravate SAH-induced cerebral ischemia and subsequent brain damage, inverse NVC may represent a novel therapeutic target after SAH.

Cerebral small vessel diseases (SVDs) are a central link between stroke and dementia—two comorbidities without specific treatments. Despite the emerging consensus that SVDs are initiated in the endothelium, the early mechanisms remain largely unknown. Deficits in on-demand delivery of blood to active brain regions (functional hyperemia) are early manifestations of the underlying pathogenesis. The capillary endothelial cell strong inward-rectifier K+ channel Kir2.1, which senses neuronal activity and initiates a propagating electrical signal that dilates upstream arterioles, is a cornerstone of functional hyperemia. Here, using a genetic SVD mouse model, we show that impaired functional hyperemia is caused by diminished Kir2.1 channel activity. We link Kir2.1 deactivation to depletion of phosphatidylinositol 4,5-bisphosphate (PIP2), a membrane phospholipid essential for Kir2.1 activity. Systemic injection of soluble PIP2 rapidly restored functional hyperemia in SVD mice, suggesting a possible strategy for rescuing functional hyperemia in brain disorders in which blood flow is disturbed.

Intracellular Ca2+ release events ('Ca 2+ sparks') and transient activation of large-conductance Ca 2+ -activated potassium (BK) channels represent an important vasodilator pathway in the cerebral vasculature. Considering the frequent occurrence of cerebral artery constriction after subarachnoid hemorrhage (SAH), our objective was to determine whether Ca 2+ spark and BK channel activity were reduced in cerebral artery myocytes from SAH model rabbits. Using laser scanning confocal microscopy, we observed B50% reduction in Ca 2+ spark activity, reflecting a decrease in the number of functional Ca 2+ spark discharge sites. Patch-clamp electrophysiology showed a similar reduction in Ca 2+ spark-induced transient BK currents, without change in BK channel density or single-channel properties. Consistent with a reduction in active Ca 2+ spark sites, quantitative real-time PCR and western blotting revealed decreased expression of ryanodine receptor type 2 (RyR-2) and increased expression of the RyR-2-stabilizing protein, FKBP12.6, in the cerebral arteries from SAH animals. Furthermore, inhibitors of Ca 2+ sparks (ryanodine) or BK channels (paxilline) constricted arteries from control, but not from SAH animals. This study shows that SAH-induced decreased subcellular Ca 2+ signaling events disable BK channel activity, leading to cerebral artery constriction. This phenomenon may contribute to decreased cerebral blood flow and poor outcome after aneurysmal SAH. Keywords: cerebral aneurysm; FKBP12.6; potassium channels; ryanodine receptors; vascular smooth muscle; vasospasm IntroductionCerebral aneurysm rupture and the ensuing subarachnoid hemorrhage (SAH) has an enormous impact on individuals and society, with 30-day mortality rates approaching 50% and the majority of survivors left with moderate-to-severe disability (Hop et al, 1997). For decades, 'angiographically defined' cerebral vasospasm of conduit arteries ( > 1 mm in diameter) has been considered to be the major contributor to death and disability in SAH patients surviving the initial intracranial bleed. However, recent evidence indicates that factors other than large-artery vasospasm contribute to SAH-induced pathologies (Macdonald et al, 2007). Additional factors contributing to the deleterious consequences of aneurysmal SAH may include global transient ischemia, early brain injury, disruption of the bloodbrain barrier, and activation of inflammatory pathways (Ostrowski et al, 2006;Prunell et al, 2005). It has now been realized that SAH may also impact small-diameter arteries and arterioles, i.e., those involved in the autoregulation of cerebral blood flow (Hattingen et al, 2008;Ishiguro et al, 2002;Ohkuma et al, 2000).In resistance arteries from healthy animals, vasoconstrictor stimuli such as physiologic increases in intravascular pressure lead to smooth muscle membrane potential depolarization, increased voltage-dependent Ca 2+ channel (VDCC) activity, and elevated global cytosolic calcium (Knot and Nelson, 1998 , the NIH (R01 HL078983, R01 HL078983-05S1, R01 ...