The goal of this work is to elucidate the cellular and molecular mechanisms that modulate alpha-1 adrenergic receptor (alpha-1 AR) mediated responses in vascular smooth muscle cells. We will focus on mechanisms that are likely to regulate 1) receptor number and cellular distribution (synthesis, degradation, cellular trafficking and post- translational modification), and 2) receptor coupling to second messenger pathways.
Specific Aim I elucidates the characteristics and mechanisms of the cellular trafficking of alpha-1 AR (internalization, externalization, degradation), and the effects of second messengers and agonist on these processes.
Specific Aim II evaluates the functional consequences of alpha-1 AR glycosylation by determining the effects of glycosidase and glycosylation inhibitors on agonist-receptor interactions, the ability to activate second messenger systems, and cellular receptor trafficking.
Specific Aim III assesses the mechanism of alpha-1 AR mRNA regulation (transcriptional, post-transcriptional, protein synthesis dependence), and the role of ionic and chemical second messengers (Ca++, cAMP, PK-C, pH) in regulating alpha-1 AR mRNA levels.
Specific Aim I V addresses the mechanism of acute agonist-induced desensitization (the roles of PK-C and receptor sequestration) and evaluates the hypothesis that there are two subtypes of alpha-1 AR that exhibit differential second messenger coupling, and possibly, differential regulation. Experiments will be performed in cultured vascular smooth muscle cells. Alpha-1 adrenergic receptor number, cellular distribution and agonist binding properties will be determined with [3H]-prazosin in intact, broken and fractionated cells with both equilibrium and non-equilibrium assays. The regulation of alpha-1 receptor mRNA levels will be evaluated using a DNA probe. The relationships between receptor properties (number, distribution, agonist affinity) and functional responses will be assessed in parallel experiments with measurements of Ca++ flux, phospholipid turnover (IP3, DAG, choline release) and arachidonic acid production. These studies should improve our understanding of the physiology of alpha-1 adrenergic control of vascular tone, and may provide insight into disease states that are characterized by abnormal vascular responsiveness to catecholamines.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL042539-04
Application #
3360823
Study Section
Experimental Cardiovascular Sciences Study Section (ECS)
Project Start
1989-04-01
Project End
1994-03-31
Budget Start
1992-04-01
Budget End
1993-03-31
Support Year
4
Fiscal Year
1992
Total Cost
Indirect Cost
Name
Brigham and Women's Hospital
Department
Type
DUNS #
071723621
City
Boston
State
MA
Country
United States
Zip Code
02115
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