In the CNS, astrocytes are sensory and regulatory hubs that play important roles in cerebral homeostatic processes, including matching local cerebral blood flow to neuronal metabolism (neurovascular coupling). These cells possess a highly branched network of processes that project from the soma to neuronal synapses as well as to arterioles and capillaries, where they terminate in "endfeet" that encase the blood vessels. Ca 2+ signaling within the endfoot mediates neurovascular coupling; thus, these functional microdomains control vascular tone and local perfusion in the brain. Transient receptor potential vanilloid 4 (TRPV4) channels-nonselective cation channels with considerable Ca 2+ conductance-have been identified in astrocytes, but their function is largely unknown. We sought to characterize the influence of TRPV4 channels on Ca 2+ dynamics in the astrocytic endfoot microdomain and assess their role in neurovascular coupling. We identified local TRPV4-mediated Ca 2+ oscillations in endfeet and further found that TRPV4 Ca 2+ signals are amplified and propagated by Ca 2+ -induced Ca 2+ release from inositol trisphosphate receptors (IP 3 Rs). Moreover, TRPV4-mediated Ca 2+ influx contributes to the endfoot Ca 2+ response to neuronal activation, enhancing the accompanying vasodilation. Our results identify a dynamic synergy between TRPV4 channels and IP 3 Rs in astrocyte endfeet and demonstrate that TRPV4 channels are engaged in and contribute to neurovascular coupling.calcium | parenchymal arteriole A strocytes are glial cells in the brain that are essential for the structural and functional integrity of the central nervous system. Astrocytes maintain cerebral homeostasis by acting as "switchboards," receiving and integrating communication from the surrounding microenvironment and translating that information into physiological and homeostatic responses. Numerous astrocytic projections make contact with neighboring synapses, while other projections terminate in "endfeet" that spread out and wrap around parenchymal arterioles and capillaries within the brain (1, 2). This structural orientation allows astrocytes to monitor synaptic activity in neuronal networks and mediate communication between neurons and the cerebral microcirculation.Calcium (Ca 2+ ) signaling is critical for astrocyte function. Transient increases in intracellular Ca 2+ concentration ([Ca 2+ ] i ) mediated by inositol 1,4,5-trisphosphate (IP 3 ) receptor Ca 2+ release channels (IP 3 Rs) in endoplasmic reticulum (ER) membranes drive the release of chemical transmitters like glutamate, adenosine triphosphate (ATP), and D-Serine that modulate synaptic transmission and neuronal excitability (3, 4). IP 3 Rdependent Ca 2+ signaling in astrocytes is also critical for neurovascular coupling (NVC), the process by which local cerebral blood flow (CBF) is matched to neuronal metabolism (5, 6). As neuronal activity increases, synaptically released glutamate binds to metabotropic glutamate receptors (mGluRs) on perisynaptic astrocytic projections, stimula...

Dunn KM, Renic M, Flasch AK, Harder DR, Falck J, Roman RJ. Elevated production of 20-HETE in the cerebral vasculature contributes to severity of ischemic stroke and oxidative stress in spontaneously hypertensive rats. Am J Physiol Heart Circ Physiol 295: H2455-H2465, 2008. First published October 17, 2008 doi:10.1152/ajpheart.00512.2008.-Hypertension is a major risk factor for stroke, but the factors that contribute to the increased incidence and severity of ischemic stroke in hypertension remain to be determined. 20-hydroxyeicosatetraenoic acid (20-HETE) has been reported to be a potent constrictor of cerebral arteries, and inhibitors of 20-HETE formation reduce infarct size following cerebral ischemia. The present study examined whether elevated production of 20-HETE in the cerebral vasculature could contribute to the larger infarct size previously reported after transient middle cerebral artery occlusion (MCAO) in hypertensive strains of rat [spontaneously hypertensive rat (SHR) and spontaneously hypertensive stroke-prone rat (SHRSP)]. The synthesis of 20-HETE in the cerebral vasculature of SHRSP measured by liquid chromatography-tandem mass spectrometry was about twice that seen in Wistar-Kyoto (WKY) rats. This was associated with the elevated expression of cytochrome P-450 (CYP)4A protein and CYP4A1 and CYP4A8 mRNA. Infarct volume after transient MCAO was greater in SHRSP (36 Ϯ 4% of hemisphere volume) than in SHR (19 Ϯ 5%) or WKY rats (5 Ϯ 2%). This was associated with a significantly greater reduction in regional cerebral blood flow (rCBF) in SHR and SHRSP than in WKY rats during the ischemic period (78% vs. 62%). In WKY rats, rCBF returned to 75% of control following reperfusion. In contrast, SHR and SHRSP exhibited a large (166 Ϯ 18% of baseline) and sustained (1 h) postischemic hyperperfusion. Acute blockade of the synthesis of -formamidine (HET0016; 1 mg/kg) reduced infarct size by 59% in SHR and 87% in SHRSP. HET0016 had no effect on the fall in rCBF during MCAO but eliminated the hyperemic response. HET0016 also attenuated vascular O2•Ϫ formation and restored endothelium-dependent dilation in cerebral arteries of SHRSP. These results indicate the production of 20-HETE is elevated in the cerebral vasculature of SHRSP and contributes to oxidative stress, endothelial dysfunction, and the enhanced sensitivity to ischemic stroke in this hypertensive model. 20-hydroxyeicosatetraenoic acid; middle cerebral artery occlusion; cytochrome P-450A; N-hydroxy-N'-(4-butyl-2-methylphenyl)-formamidine HYPERTENSION IS A MAJOR RISK factor for stroke, but the factors that contribute to the increased incidence and severity of ischemic stroke in hypertension remain to be determined (46). Previous studies have indicated that arachidonic acid (AA) is released into cerebrospinal fluid following cerebral ischemia (1,31,42,56). AA is converted by cytochrome P-450 (CYP) enzymes in cerebral arteries to a potent vasoconstrictor, 20-hydroxyeicosatetraenoic acid (20-HETE) (15,16,32). 20-HETE has been reported to play an important...

Calcium-sensitive potassium (K(Ca)) channels have been shown to modulate the diameter of cerebral pial arteries; however, little is known regarding their roles in controlling cerebral parenchymal arterioles (PAs). We explored the function and cellular distribution of small-conductance (SK(Ca)) and intermediate-conductance (IK(Ca)) K(Ca) channels and large-conductance K(Ca) (BK(Ca)) channels in endothelial cells (ECs) and smooth muscle cells (SMCs) of PAs. Both SK(Ca) and IK(Ca) channels conducted the outward current in isolated PA ECs (current densities, ~20 pA/pF and ~28 pA/pF at +40 mV, respectively), but these currents were not detected in PA SMCs. In contrast, BK(Ca) currents were prominent in PA SMCs (~154 pA/pF), but were undetectable in PA ECs. Pressurized PAs constricted to inhibition of SK(Ca) (~16%) and IK(Ca) (~16%) channels, but were only modestly affected by inhibition of BK(Ca) channels (~5%). Blockade of SK(Ca) and IK(Ca) channels decreased resting cortical cerebral blood flow (CBF) by ~15%. NS309 (6,7-dichloro-1H-indole-2,3-dione3-oxime), a SK(Ca)/IK(Ca) channel opener, hyperpolarized PA SMCs by ~27 mV, maximally dilated pressurized PAs, and increased CBF by ~40%. In conclusion, these data show that SK(Ca) and IK(Ca) channels in ECs profoundly modulate PA tone and CBF, whereas BK(Ca) channels in SMCs only modestly influence PA diameter.

Neuronal activity is communicated to the cerebral vasculature so that adequate perfusion of brain tissue is maintained at all levels of neuronal metabolism. An increase in neuronal activity is accompanied by vasodilation and an increase in local cerebral blood flow. This process, known as neurovascular coupling (NVC) or functional hyperemia, is essential for cerebral homeostasis and survival. Neuronal activity is encoded in astrocytic Ca 2+ signals that travel to astrocytic processes ('endfeet') encasing parenchymal arterioles within the brain. Astrocytic Ca 2+ signals cause the release of vasoactive substances to cause relaxation, and in some circumstances contraction, of the smooth muscle cells (SMCs) of parenchymal arterioles to modulate local cerebral blood flow. Activation of potassium channels in the SMCs has been proposed to mediate NVC. Here, the current state of knowledge of NVC and potassium channels in parenchymal arterioles is reviewed. (Circ J 2010; 74: 608 - 616)

Recent studies have indicated that arachidonic acid (AA) is metabolized by the cytochrome P450 4A (CYP4A) enzymes in cerebral arteries to produce 20-hydroxyeicosatetraenoic acid (20-HETE) and that this compound has effects on cerebral vascular tone that mimic those seen following subarachnoid hemorrhage (SAH). In this regard, 20-HETE is a potent constrictor of cerebral arteries that decreases the open state probability of Ca(2+)-activated K(+) channels through activation of protein kinase C (PKC). It increases the sensitivity of the contractile apparatus to Ca(2+) by activating PKC and rho kinase. The formation of 20-HETE is stimulated by angiotensin II (AII), endothelin, adenosine triphosphate (ATP) and serotonin, and inhibited by NO, CO and superoxide radicals. Inhibitors of the formation of 20-HETE block the myogenic response of cerebral arterioles to elevations in transmural pressure in vitro and autoregulation of cerebral blood flow (CBF) in vivo. 20-HETE also plays an important role in modulating the cerebral vascular responses to vasodilators (NO and CO) and vasoconstrictors (AII, endothelin, serotonin). Recent studies have indicated that the levels of 20-HETE in cerebrospinal fluid (CSF) increase in rats, dogs and human patients following SAH and that inhibitors of the synthesis of 20-HETE prevent the acute fall in CBF in rats and reverse delayed vasospasm in both dogs and rats. This review examines the evidence that an elevation in the production of 20-HETE contributes to the initial fall in CBF following SAH and the later development of delayed vasospasm.