Adenosine triphosphate (ATP) is a potent regulator of vascular tone in the cerebral circulation. This physiological agonist is released from a variety of sources in the cardiovascular system under normal and pathological conditions. When presented to the endothelium, ATP stimulates artery dilation through multiple pathways including endothelium-derived hyperpolarizing factor (EDHF) mediated mechanisms. Endothelial cell (EC) hyperpolarization plays a crucial role in EDHF mediated dilation and is a fundamental determinant of arterial tone in multiple vascular beds. While the mechanism by which ECs become hyperpolarized by agonists is still poorly defined, several lines of evidence indicate a significant role for activation of endothelial intermediate-conductance KCa (IKCa) channels. IKCa channels are primarily regulated by cytosolic Ca2+ concentration, but the source of the activating Ca2+ is not known. We propose that ATP stimulates Ca2+ influx through endothelial transient receptor potential (TRP) channels which promotes IKCa channel activation with subsequent EC hyperpolarization and artery dilation. Specifically, we propose to:
Aim 1 : Define the role of TRP channels in ATP signaling in cerebrovascular endothelial cells. ATP promotes endothelial Ca2+ influx and subsequent dilation via NO and EDHF dependent mechanisms. In this specific aim we will determine if ATP activates TRP channels to promote Ca2+ influx in cerebrovascular endothelial cells. We will identify the TRP channels expressed in ECs (RT-PCR and immunohistochemistry), measure [Ca2+]i and Ca2+ influx in response to ATP in freshly isolated middle cerebral artery (MCA) ECs and in the ECs of pressurized MCA (Fura 2 dye), and demonstrate the role of the identified TRP channels by pharmacological and RNA silencing techniques (organ culture with siRNA).
Aim 2 : Elucidate the role of TRP channels in IKCa channel activation, EC hyperpolarization, and EDHF-mediated dilation in cerebral arteries. EDHF-mediated dilation of cerebral arteries requires EC Ca2+ influx, IKCa channel activation, and EC hyperpolarization. We will determine the role of Ca2+ influx through TRP channels on IKCa channel activation and EC hyperpolarization. Specifically, we will demonstrate that Ca2+ influx via TRP channels is critical for activation of IKCa channels and subsequent EC hyperpolarization using techniques to measure membrane potential and IKCa channel activation (whole cell patch clamp) in ECs from organ cultured MCA. We will also demonstrate that Ca2+ influx via TRP channels is critical for EDHF-mediated dilation by measuring ATP stimulated dilation in intact organ cultured MCA. These studies should lead to novel therapeutic strategies for controlling blood flow in the brain by defining the role of specific TRP channels in the regulation of EC Ca2+ concentration, EC hyperpolarization, and EDHF-mediated dilation. Adenosine triphosphate (ATP) is released into the cerebral circulation and acts on endothelial cells within cerebral arteries to control blood flow in the brain. However, we do not presently understand the mechanism by which ATP controls this endothelial cell function. These studies will define this mechanism of blood flow control and thus provide novel therapeutic strategies for regulating cerebral blood flow in health and disease states.
|Feketa, Viktor V; Zhang, Yi; Cao, Zhijuan et al. (2014) Transient receptor potential melastatin 8 channel inhibition potentiates the hypothermic response to transient receptor potential vanilloid 1 activation in the conscious mouse. Crit Care Med 42:e355-63|
|Cao, Zhijuan; Balasubramanian, Adithya; Marrelli, Sean P (2014) Pharmacologically induced hypothermia via TRPV1 channel agonism provides neuroprotection following ischemic stroke when initiated 90 min after reperfusion. Am J Physiol Regul Integr Comp Physiol 306:R149-56|
|Kochukov, Mikhail Y; Balasubramanian, Adithya; Abramowitz, Joel et al. (2014) Activation of endothelial transient receptor potential C3 channel is required for small conductance calcium-activated potassium channel activation and sustained endothelial hyperpolarization and vasodilation of cerebral artery. J Am Heart Assoc 3:|
|Lloyd, Eric E; Pandit, Lavannya M; Crossland, Randy F et al. (2013) Endothelium-dependent relaxations in the aorta from K(2p)6.1 knockout mice. Am J Physiol Regul Integr Comp Physiol 305:R60-7|
|Kochukov, M Y; Balasubramanian, A; Noel, R C et al. (2013) Role of TRPC1 and TRPC3 channels in contraction and relaxation of mouse thoracic aorta. J Vasc Res 50:11-20|
|Crossland, Randy F; Durgan, David J; Lloyd, Eric E et al. (2013) A new rodent model for obstructive sleep apnea: effects on ATP-mediated dilations in cerebral arteries. Am J Physiol Regul Integr Comp Physiol 305:R334-42|
|Feketa, Viktor V; Balasubramanian, Adithya; Flores, Christopher M et al. (2013) Shivering and tachycardic responses to external cooling in mice are substantially suppressed by TRPV1 activation but not by TRPM8 inhibition. Am J Physiol Regul Integr Comp Physiol 305:R1040-50|
|Lloyd, Eric E; Crossland, Randy F; Phillips, Sharon C et al. (2011) Disruption of K(2P)6.1 produces vascular dysfunction and hypertension in mice. Hypertension 58:672-8|
|Namiranian, Khodadad; Lloyd, Eric E; Crossland, Randy F et al. (2010) Cerebrovascular responses in mice deficient in the potassium channel, TREK-1. Am J Physiol Regul Integr Comp Physiol 299:R461-9|
|Lloyd, Eric E; Marrelli, Sean P; Bryan Jr, Robert M (2009) cGMP does not activate two-pore domain K+ channels in cerebrovascular smooth muscle. Am J Physiol Heart Circ Physiol 296:H1774-80|
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