Combining In Vivo Two-Photon and Laser Speckle Microscopy With the Ex Vivo Capillary-Parenchymal Arteriole Preparation as a Novel Approach to Study Neurovascular Coupling.

Objective: Cerebral blood flow (CBF) decline is increasingly recognized as an area of importance for targeting neurodegenerative disorders, yet full understanding of the mechanisms that underlie CBF changes are lacking. Animal models are crucial for expanding our knowledge as methods for studying global CBF and neurovascular coupling in humans are limited and require expensive specialized scanners.

Methods: Use of appropriate animal models can increase our understanding of cerebrovascular function, so we have combined chronic cranial windows with in vivo two-photon and laser speckle microscopy and ex vivo capillary-parenchymal arteriole (CaPA) preparations.

A multi-disciplinary commentary on preclinical research to investigate vascular contributions to dementia.

Although dementia research has been dominated by Alzheimer's disease (AD), most dementia in older people is now recognised to be due to mixed pathologies, usually combining vascular and AD brain pathology. Vascular cognitive impairment (VCI), which encompasses vascular dementia (VaD) is the second most common type of dementia. Models of VCI have been delayed by limited understanding of the underlying aetiology and pathogenesis.

Uncoupling of Ca sparks from BK channels in cerebral arteries underlies hypoperfusion in hypertension-induced vascular dementia.

The deficit in cerebral blood flow (CBF) seen in patients with hypertension-induced vascular dementia is increasingly viewed as a therapeutic target for disease-modifying therapy. Progress is limited, however, due to uncertainty surrounding the mechanisms through which elevated blood pressure reduces CBF. To investigate this, we used the BPH/2 mouse, a polygenic model of hypertension.

Functionally linked potassium channel activity in cerebral endothelial and smooth muscle cells is compromised in Alzheimer's disease.

The brain microcirculation is increasingly viewed as a potential target for disease-modifying drugs in the treatment of Alzheimer's disease patients, reflecting a growing appreciation of evidence that cerebral blood flow is compromised in such patients. However, the pathogenic mechanisms in brain resistance arteries underlying blood flow defects have not yet been elucidated. Here we probed the roles of principal vasodilatory pathways in cerebral arteries using the APP23 mouse model of Alzheimer's disease, in which amyloid precursor protein is increased approximately sevenfold, leading to neuritic plaques and cerebrovascular accumulation of amyloid-β similar to those in patients with Alzheimer's disease.

Cardiovascular comorbidities, inflammation, and cerebral small vessel disease.

Cerebral small vessel disease (cSVD) is the most common cause of vascular cognitive impairment and affects all levels of the brain's vasculature. Features include diverse structural and functional changes affecting small arteries and capillaries that lead to a decline in cerebral perfusion. Due to an ageing population, incidence of cSVD is continually rising.

Kynurenine Relaxes Arteries of Normotensive Women and Those With Preeclampsia.

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TRPML1 channels initiate Ca sparks in vascular smooth muscle cells.

TRPML1 (transient receptor potential mucolipin 1) is a Ca-permeable, nonselective cation channel localized to the membranes of endosomes and lysosomes and is not present or functional on the plasma membrane. Ca released from endosomes and lysosomes into the cytosol through TRPML1 channels is vital for trafficking, acidification, and other basic functions of these organelles. Here, we investigated the function of TRPML1 channels in fully differentiated contractile vascular smooth muscle cells (SMCs).

Nanoscale coupling of junctophilin-2 and ryanodine receptors regulates vascular smooth muscle cell contractility.

Junctophilin proteins maintain close contacts between the endoplasmic/sarcoplasmic reticulum (ER/SR) and the plasma membrane in many types of cells, as typified by junctophilin-2 (JPH2), which is necessary for the formation of the cardiac dyad. Here, we report that JPH2 is the most abundant junctophilin isotype in native smooth muscle cells (SMCs) isolated from cerebral arteries and that acute knockdown diminishes the area of sites of interaction between the SR and plasma membrane. Superresolution microscopy revealed nanometer-scale colocalization of JPH2 clusters with type 2 ryanodine receptor (RyR2) clusters near the cell surface.

Nanoscale remodeling of ryanodine receptor cluster size underlies cerebral microvascular dysfunction in Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) results from mutations in the gene encoding dystrophin which lead to impaired function of skeletal and cardiac muscle, but little is known about the effects of the disease on vascular smooth muscle cells (SMCs). Here we used the mouse model to study the effects of mutant dystrophin on the regulation of cerebral artery and arteriole SMC contractility, focusing on an important Ca-signaling pathway composed of type 2 ryanodine receptors (RyR2s) on the sarcoplasmic reticulum (SR) and large-conductance Ca-activated K (BK) channels on the plasma membrane. Nanoscale superresolution image analysis revealed that RyR2 and BKα were organized into discrete clusters, and that the mean size of RyR2 clusters that colocalized with BKα was larger in SMCs from mice (∼62 RyR2 monomers) than in controls (∼40 RyR2 monomers).

Microtubule structures underlying the sarcoplasmic reticulum support peripheral coupling sites to regulate smooth muscle contractility.

Junctional membrane complexes facilitate excitation-contraction coupling in skeletal and cardiac muscle cells by forming subcellular invaginations that maintain close (≤20 nm) proximity of ryanodine receptors (RyRs) on the sarcoplasmic reticulum (SR) with voltage-dependent Ca channels in the plasma membrane. In fully differentiated smooth muscle cells, junctional membrane complexes occur as distributed sites of peripheral coupling. We investigated the role of the cytoskeleton in maintaining peripheral coupling and associated Ca signaling networks within native smooth muscle cells of mouse and rat cerebral arteries.

The angiotensin II receptor type 1b is the primary sensor of intraluminal pressure in cerebral artery smooth muscle cells.

The angiotensin II receptor type 1b (AT R ) is the primary sensor of intraluminal pressure in cerebral arteries. Pressure or membrane-stretch induced stimulation of AT R activates the TRPM4 channel and results in inward transient cation currents that depolarize smooth muscle cells, leading to vasoconstriction. Activation of either AT R or AT R with angiotensin II stimulates TRPM4 currents in cerebral artery myocytes and vasoconstriction of cerebral arteries.

Unitary TRPV3 channel Ca2+ influx events elicit endothelium-dependent dilation of cerebral parenchymal arterioles.

Cerebral parenchymal arterioles (PA) regulate blood flow between pial arteries on the surface of the brain and the deeper microcirculation. Regulation of PA contractility differs from that of pial arteries and is not completely understood. Here, we investigated the hypothesis that the Ca(2+) permeable vanilloid transient receptor potential (TRPV) channel TRPV3 can mediate endothelium-dependent dilation of cerebral PA.

Molecular and functional significance of Ca(2+)-activated Cl(-) channels in pulmonary arterial smooth muscle.

Increased peripheral resistance of small distal pulmonary arteries is a hallmark signature of pulmonary hypertension (PH) and is believed to be the consequence of enhanced vasoconstriction to agonists, thickening of the arterial wall due to remodeling, and increased thrombosis. The elevation in arterial tone in PH is attributable, at least in part, to smooth muscle cells of PH patients being more depolarized and displaying higher intracellular Ca(2+) levels than cells from normal subjects. It is now clear that downregulation of voltage-dependent K(+) channels (e.

Potent vasorelaxant activity of the TMEM16A inhibitor T16A(inh) -A01.

Background And Purpose: T16A(inh) -A01 is a recently identified inhibitor of the calcium-activated chloride channel TMEM16A. The aim of this study was to test the efficacy of T16A(inh) -A01 for inhibition of calcium-activated chloride channels in vascular smooth muscle and consequent effects on vascular tone.

Experimental Approach: Single channel and whole cell patch clamp was performed on single smooth muscle cells from rabbit pulmonary artery and mouse thoracic aorta.