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Intraluminal pressure stimulates MAPK phosphorylation in arterioles: temporal dissociation from myogenic contractile
Brian E Spurrell1, Timothy V Murphy, Michael A Hill
1Microvascular Biology Group, School of Medical Sciences, RMIT University, Plenty Rd., Bundoora, Victoria 3083, Australia.
Abstract:
Members of the MAPK family of enzymes, p42/44 and p38, have been implicated in both the regulation of contractile function and growth responses in vascular smooth muscle. We determined whether such kinases are activated during the arteriolar myogenic response after increases in intraluminal pressure. Particular emphasis was placed on temporal aspects of activation to determine whether such phosphorylation events parallel the known time course for myogenic contraction. Experiments used single cannulated arterioles isolated from the cremaster muscle of rats with some vessels loaded with the fluorescent Ca2+-sensitive dye fura 2 (2 microM). The p42/44 inhibitor PD-98059 (50 microM) caused vasodilation but did not prevent pressure-induced myogenic constriction. The vasodilator response was accompanied by decreased smooth muscle intracellular Ca2+. Western blotting revealed a significant increase in the level of phosphorylation of p42/44 15 min after the application of a 30- to 100-mmHg pressure step. Phosphorylation of p42/44 was a late event that appeared to be temporally dissociated from contraction, which was complete within 1-5 min. EGF (80 nM) caused marked phosphorylation of p42/44 but only acted as a weak vasoconstrictor. The p38 inhibitor SB-203580 (10 microM) did not alter baseline diameter, nor did it prevent myogenic vasoconstriction. Consistent with these observations, SB-203580 did not cause a measurable change in intracellular Ca2+. The results demonstrate activation of the p42/44 class of MAPK resulting from increased transmural pressure. Such activation is, however, dissociated from the acute pressure-induced vasoconstrictor response in terms of time course and may represent the activation of compensatory, but parallel, pathways, including those related to growth and remodeling.
Insights
Mitogen-activated protein kinases (MAPK) p42/44 are activated by increased pressure in arterioles, but this response is delayed and separate from the rapid myogenic constriction. This suggests parallel pathways involved in vascular smooth muscle adaptation.
Area of Science:
- Physiology
- Molecular Biology
- Vascular Biology
Background:
- Mitogen-activated protein kinase (MAPK) family members, including p42/44 and p38, are involved in vascular smooth muscle (VSM) contractile and growth regulation.
- The myogenic response is a critical VSM function that maintains blood flow homeostasis by constricting in response to increased intraluminal pressure.
Purpose of the Study:
- To investigate the activation patterns of p42/44 and p38 MAPK during the arteriolar myogenic response.
- To determine if MAPK activation temporally correlates with the onset and progression of myogenic contraction.
Main Methods:
- Single cannulated rat cremaster arterioles were pressurized, and some were loaded with fura 2 for intracellular calcium (Ca2+) measurement.
- Western blotting was used to assess the phosphorylation levels of p42/44 and p38 MAPK.
- Pharmacological inhibitors (PD-98059 for p42/44, SB-203580 for p38) were used to probe kinase function.
Main Results:
- Increased intraluminal pressure significantly increased p42/44 MAPK phosphorylation starting at 15 minutes, which is temporally dissociated from the rapid (1-5 min) myogenic contraction.
- Inhibition of p42/44 MAPK (PD-98059) caused vasodilation and reduced intracellular Ca2+ but did not abolish myogenic constriction.
- Inhibition of p38 MAPK (SB-203580) had no effect on myogenic tone or intracellular Ca2+.
- Epidermal growth factor (EGF) induced p42/44 phosphorylation but only weak vasoconstriction.
Conclusions:
- The p42/44 MAPK pathway is activated by increased transmural pressure in arterioles.
- p42/44 MAPK activation is not directly responsible for the acute myogenic vasoconstriction, suggesting its role in parallel pathways.
- These parallel pathways may be involved in VSM growth and remodeling responses to pressure changes.