Episodic, moderate-to-heavy ethyl alcohol (ethanol) intake is a primary form of excessive alcohol consumption in the United States. Episodic drinking that results in blood alcohol levels of 35-80 mM (0.16- 0.37%) is associated with an increased risk for cerebral ischemia, stroke, and death from ischemic stroke. Cerebral ischemia may arise from or be exacerbated by enhanced constriction of cerebral arteries. Using a rat model, we previously showed that ethanol-induced cerebral artery constriction (ECAC) is caused by ethanol inhibition of calcium/voltage-gated potassium channels of large conductance (BK) in vascular smooth muscle. Moreover, ethanol-induced inhibition of BK channels at physiological conditions can be observed only in the presence of BK beta1 subunits, which are abundant in vascular smooth muscle. Factors that control BK beta1 subunit-mediated ECAC are largely unknown. Cholesterol is a characteristic constituent of Western diets. We have recently identified cholesterol as a critical nutritional regulator of ECAC, with dietary cholesterol protecting against ECAC. However, clinical applicability of our finding is limited by several fundamental questions that remain unanswered. In the current proposal, we will answer these questions by testing related, yet independently testable, hypotheses (Specific Aims 1-3): 1) define cholesterol levels both in circulation and cerebral artery tissue that represent a window of vulnerability for ethanol-induced inhibition of vascular smooth muscle BK channels and resulting ECAC, 2) determine the effect of statin therapy on ECAC and identify the underlying mechanism(s) by which statin therapy affects ECAC, 3) identify distinct amino acids within the BK channel beta1 subunit that disable ethanol- induced BK channel inhibition and results in ECAC in the presence of an elevated cholesterol level that exceeds the window of vulnerability. In the course of the study, we will use a combination of techniques that range from manipulating cholesterol levels in vivo (using a high-cholesterol diet and statin therapy) to constructing mutated BK beta1 subunits to ablate cholesterol-driven protection against ECAC. Our work will, for the first time, determine particular cholesterol levels in the blood that serve s markers of vulnerability to ECAC. Moreover, we will, for the first time, establish mechanism(s) by which statin therapy affects ECAC. Finally, we will identify distinct amino acids within the BK beta1 subunit that are responsible for cholesterol control over ethanol-induced BK channel inhibition and ECAC. The role of these amino acids will be validated at the organ level after we permeabilize cerebral arteries of beta1 knockout mice with DNA of newly engineered beta1 subunits. Our work may ultimately lead to improved preventive practices against ECAC and will open a venue for developing novel therapeutics to counteract ethanol-induced BK channel inhibition and the resulting constriction of cerebral arteries.

Public Health Relevance

Cholesterol is a characteristic ingredient in Western diet and critical regulator of alcohol effect on cerebral artery diameter. We aim to determine blood cholesterol levels that protect against alcohol-induced cerebral artery constriction, to characterize for the first time the effect of statin therapy on cerebrovascular effect of alcohol, and to identify molecular bases for cholesterol control of alcohol effect on cerebral artery. Our work will provide tool for identification of individual's vulnerability for alcohol-driven cerebrovascular pathology and will open a venue for rational drug design to combat deleterious effect of alcohol on cerebral circulation.

National Institute of Health (NIH)
National Institute on Alcohol Abuse and Alcoholism (NIAAA)
Research Project (R01)
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Neurotoxicology and Alcohol Study Section (NAL)
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Orosz, Andras
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University of Tennessee Health Science Center
Schools of Medicine
United States
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North, Kelsey; Bisen, Shivantika; Dopico, Alex M et al. (2018) Tyrosine 450 in the Voltage- and Calcium-Gated Potassium Channel of Large Conductance Channel Pore-Forming (slo1) Subunit Mediates Cholesterol Protection against Alcohol-Induced Constriction of Cerebral Arteries. J Pharmacol Exp Ther 367:234-244
Dopico, Alex M; Bukiya, Anna N; Jaggar, Jonathan H (2018) Calcium- and voltage-gated BK channels in vascular smooth muscle. Pflugers Arch 470:1271-1289
Bukiya, Anna N; Dopico, Alex M (2018) Fetal Cerebral Circulation as Target of Maternal Alcohol Consumption. Alcohol Clin Exp Res 42:1006-1018
Simakova, Maria N; Bisen, Shivantika; Dopico, Alex M et al. (2017) Statin therapy exacerbates alcohol-induced constriction of cerebral arteries via modulation of ethanol-induced BK channel inhibition in vascular smooth muscle. Biochem Pharmacol 145:81-93
Dopico, Alex M; Bukiya, Anna N (2017) Regulation of Ca2+-Sensitive K+ Channels by Cholesterol and Bile Acids via Distinct Channel Subunits and Sites. Curr Top Membr 80:53-93
Dopico, Alex M; Bukiya, Anna N; Bettinger, Jill C (2017) Voltage-Sensitive Potassium Channels of the BK Type and Their Coding Genes Are Alcohol Targets in Neurons. Handb Exp Pharmacol :
Bukiya, Anna N; Dopico, Alejandro M (2017) Common structural features of cholesterol binding sites in crystallized soluble proteins. J Lipid Res 58:1044-1054
Dopico, A M; Bukiya, A N; Kuntamallappanavar, G et al. (2016) Modulation of BK Channels by Ethanol. Int Rev Neurobiol 128:239-79
Bisen, Shivantika; Seleverstov, Olga; Belani, Jitendra et al. (2016) Distinct mechanisms underlying cholesterol protection against alcohol-induced BK channel inhibition and resulting vasoconstriction. Biochim Biophys Acta 1861:1756-1766