Angiotensin II-mediated hypertension in the rat increases vascular superoxide production via membrane NADH/NADPH oxidase activation. Contribution to alterations of vasomotor tone.
Objective: Test the hypothesis that angiotensin II-induced hypertension is associated with increased vascular superoxide production and characterize the oxidase involved in this process
Materials & Equipment Checklist
11 items
Gather these items before starting the experiment. Check off items as you prepare.
Equipment2
Not specified • Not specified • Not specified • Not mentioned
As an Amazon Associate, we earn from qualifying purchases. Product links help support this free resource.
View Abstract
We tested the hypothesis that angiotensin II-induced hypertension is associated with an increase in vascular .O2- production, and characterized the oxidase involved in this process. Infusion of angiotensin II (0.7 mg/kg per d) increased systolic blood pressure and doubled vascular .O2- production (assessed by lucigenin chemiluminescence), predominantly from the vascular media. NE infusion (2.75 mg/kg per d) produced a similar degree of hypertension, but did not increase vascular .O2- production. Studies using various enzyme inhibitors and vascular homogenates suggested that the predominant source of .O2- activated by angiotensin II infusion is an NADH/NADPH-dependent, membrane-bound oxidase. Angiotensin II-, but not NE-, induced hypertension was associated with impaired relaxations to acetylcholine, the calcium ionophore A23187, and nitroglycerin. These relaxations were variably corrected by treatment of vessels with liposome-encapsulated superoxide dismutase. When Losartan was administered concomitantly with angiotensin II, vascular .O2- production and relaxations were normalized, demonstrating a role for the angiotensin type-1 receptor in these processes. We conclude that forms of hypertension associated with elevated circulating levels of angiotensin II may have unique vascular effects not shared by other forms of hypertension because they increase vascular smooth muscle .O2- production via NADH/NADPH oxidase activation.
Protocol Steps
1
Angiotensin II infusion
Infuse angiotensin II systemically into rats at a dose of 0.7 mg/kg per day to induce hypertension
Not specifiedNot specified
Note: This treatment increases systolic blood pressure and doubles vascular superoxide production
View evidence from paper
“Infusion of angiotensin II (0.7 mg/kg per d) increased systolic blood pressure and doubled vascular .O2- production”
2
Norepinephrine infusion control
Infuse norepinephrine systemically into control rats at a dose of 2.75 mg/kg per day to produce similar degree of hypertension
Not specifiedNot specified
Note: This control treatment produces hypertension but does not increase vascular superoxide production
View evidence from paper
“NE infusion (2.75 mg/kg per d) produced a similar degree of hypertension, but did not increase vascular .O2- production”
3
Measure vascular superoxide production
Assess vascular superoxide production using lucigenin chemiluminescence in vascular tissue samples
Not specifiedNot specified
Note: Superoxide production is predominantly from the vascular media
View evidence from paper
“doubled vascular .O2- production (assessed by lucigenin chemiluminescence), predominantly from the vascular media”
4
Characterize oxidase source
Use various enzyme inhibitors and vascular homogenates to identify the source of superoxide production
Not specifiedNot specified
Note: Studies identified NADH/NADPH-dependent, membrane-bound oxidase as the predominant source
View evidence from paper
“Studies using various enzyme inhibitors and vascular homogenates suggested that the predominant source of .O2- activated by angiotensin II infusion is an NADH/NADPH-dependent, membrane-bound oxidase”
5
Assess vascular relaxation responses
Test vascular relaxation responses to acetylcholine, calcium ionophore A23187, and nitroglycerin
Not specifiedNot specified
Note: Angiotensin II-induced hypertension impairs relaxations to these agents, while NE-induced hypertension does not
View evidence from paper
“Angiotensin II-, but not NE-, induced hypertension was associated with impaired relaxations to acetylcholine, the calcium ionophore A23187, and nitroglycerin”
6
Test superoxide dismutase treatment
Treat vessels with liposome-encapsulated superoxide dismutase to determine if it corrects impaired relaxations
Not specifiedNot specified
Note: Treatment variably corrected the impaired relaxations
View evidence from paper
“These relaxations were variably corrected by treatment of vessels with liposome-encapsulated superoxide dismutase”
7
Losartan co-administration
Administer Losartan (angiotensin type-1 receptor antagonist) concomitantly with angiotensin II infusion
Not specifiedNot specified
Note: Losartan normalizes vascular superoxide production and relaxations, demonstrating a role for angiotensin type-1 receptor
View evidence from paper
“When Losartan was administered concomitantly with angiotensin II, vascular .O2- production and relaxations were normalized, demonstrating a role for the angiotensin type-1 receptor”