Abdallah Hayar
Sung Rhee
Shaker-type, voltage-gated K+ (KV1) channels are an important determinant of the resting membrane potential and diameter of small cerebral arteries. During hypertension, KV1 channel-mediated dilation appears to be blunted and is postulated to increase myogenicity in the cerebral circulation. However, little is known about the mechanisms that regulate the expression of KV1 channels at the plasma membrane of cerebral vascular smooth muscle cells (cVSMCs). In this regard, we recently identified scaffolding proteins including PSD95 (postsynaptic density 95) in rat cVSMCs that have never been described. PSD95 is a wellcharacterized scaffolding protein in neurons with more than 50 known binding partners that can facilitate macromolecular signaling between ion channels and receptors. Subsequently we determined that KV1 channels associate with the PSD95 scaffold in cVSMCs, and that PSD95 is required for the normal expression and dilator function of KV1 channels in small cerebral arteries. Finally, we have evidence that the β1 adrenergic receptor (β1AR) – another known binding partner of the PSD95 scaffold – activates a KV1 channel-mediated dilator pathway. Thus, we envision that PSD95 enables the efficient coupling of the β1AR signaling pathway to KV1 channels in cVSMCs and we have designed experiments to characterize the impact of this novel PSD95 complex on cerebrovascular reactivity. Based on our early findings, we hypothesize that: β1AR and the KV1 channels form a macromolecular vasodilator complex on a PSD95 scaffold in the rat cerebral circulation. We further propose that the down-regulation of cerebrovascular KV1 channels during hypertension disrupts the PSD95 scaffold resulting in a synchronized loss of the β1AR-KV1 signaling pathway and a vasodilator defect. These hypotheses will be tested using co-immunoprecipitation and confocal microscopy to discern protein interactions in small cerebral arteries. The physiological impact of siRNA knockdown of PSD95 or KV1 channels in cerebral arteries in vitro and in vivo will be evaluated using patch-clamp electrophysiology, microvessel reactivity assays, and intravital microscopy. The findings of this project will identify for the first time a vasodilator complex in vascular smooth muscle that is regulated by scaffolding proteins, and will set the stage for further studies to understand how ion channels are localized with their signaling partners in cVSMCs.
Project 2: An ultra-fast imaging
method for monitoring blood flow in the cerebral microcirculation
The etiology of brain hemorrhage is multifactorial and can be induced by extreme alterations in many physiological parameters that influence cerebral blood flow (CBF) and the capacity for cerebral autoregulation, such as blood gases, arterial blood pressure, body temperature, the autonomic nervous system, and neuronal activity. Using animal research, this exploratory project aims at a better understanding of the physiological factors that influence CBF and cerebral autoregulation and the different factors leading to cerebral hemorrhage. The main goal of this project is to better understand the mechanisms that would favor or trigger cerebral hemorrhage. We hypothesize that small capillaries are the most structurally vulnerable part of the cerebral circulation and may rupture in extreme physiological conditions leading to cerebral hemorrhage. We think that investigating CBF at the level of single capillaries using an ultra-fast imaging method would provide new insight into how small microvessels react in response to different physiological stressors such as hypercapnia, high blood pressure and excessive neuronal excitability. Optical imaging techniques, nowadays, have become powerful tools for investigating CBF. Our novel method consists of imaging and quantifying cerebral microcirculation in rat olfactory bulb capillaries at 250-2000 frames/sec. Unlike laser line-scanning microscopy which is relatively slow to acquire full frame images, our method will allow to perform live imaging of red blood cell traffic at very high frame rate. We will test whether weakening the wall of blood vessels with ultraviolet laser irradiation in addition to inducing extreme physiological changes can deleteriously alter the microcirculation and result in cerebral hemorrhage. This project also aims at finding the optimal neuroprotective conditions that may prevent the rupture of vulnerable microvessels during extreme physiological conditions. By seeking a better understanding of the mechanisms leading to brain hemorrhage, the results of this research project will allow us to develop new strategies in the prevention and treatment of cerebral hemorrhage.
The etiology of brain hemorrhage is multifactorial and can be induced by extreme alterations in many physiological parameters that influence cerebral blood flow (CBF) and the capacity for cerebral autoregulation, such as blood gases, arterial blood pressure, body temperature, the autonomic nervous system, and neuronal activity. Using animal research, this exploratory project aims at a better understanding of the physiological factors that influence CBF and cerebral autoregulation and the different factors leading to cerebral hemorrhage. The main goal of this project is to better understand the mechanisms that would favor or trigger cerebral hemorrhage. We hypothesize that small capillaries are the most structurally vulnerable part of the cerebral circulation and may rupture in extreme physiological conditions leading to cerebral hemorrhage. We think that investigating CBF at the level of single capillaries using an ultra-fast imaging method would provide new insight into how small microvessels react in response to different physiological stressors such as hypercapnia, high blood pressure and excessive neuronal excitability. Optical imaging techniques, nowadays, have become powerful tools for investigating CBF. Our novel method consists of imaging and quantifying cerebral microcirculation in rat olfactory bulb capillaries at 250-2000 frames/sec. Unlike laser line-scanning microscopy which is relatively slow to acquire full frame images, our method will allow to perform live imaging of red blood cell traffic at very high frame rate. We will test whether weakening the wall of blood vessels with ultraviolet laser irradiation in addition to inducing extreme physiological changes can deleteriously alter the microcirculation and result in cerebral hemorrhage. This project also aims at finding the optimal neuroprotective conditions that may prevent the rupture of vulnerable microvessels during extreme physiological conditions. By seeking a better understanding of the mechanisms leading to brain hemorrhage, the results of this research project will allow us to develop new strategies in the prevention and treatment of cerebral hemorrhage.