Role of myosin light chain kinase in regulation of basal blood pressure and maintenance of salt-induced hypertension

Abstract

Vascular tone, an important determinant of systemic vascular resistance and thus blood pressure, is affected by vascular smooth muscle (VSM) contraction. Key signaling pathways for VSM contraction converge on phosphorylation of the regulatory light chain (RLC) of smooth muscle myosin. This phosphorylation is mediated by Ca(2+)/calmodulin-dependent myosin light chain kinase (MLCK) but Ca(2+)-independent kinases may also contribute, particularly in sustained contractions. Signaling through MLCK has been indirectly implicated in maintenance of basal blood pressure, whereas signaling through RhoA has been implicated in salt-induced hypertension. In this report, we analyzed mice with smooth muscle-specific knockout of MLCK. Mesenteric artery segments isolated from smooth muscle-specific MLCK knockout mice (MLCK(SMKO)) had a significantly reduced contractile response to KCl and vasoconstrictors. The kinase knockout also markedly reduced RLC phosphorylation and developed force. We suggest that MLCK and its phosphorylation of RLC are required for tonic VSM contraction. MLCK(SMKO) mice exhibit significantly lower basal blood pressure and weaker responses to vasopressors. The elevated blood pressure in salt-induced hypertension is reduced below normotensive levels after MLCK attenuation. These results suggest that MLCK is necessary for both physiological and pathological blood pressure. MLCK(SMKO) mice may be a useful model of vascular failure and hypotension.

Myosin Phosphatase Target Subunit 1 (MYPT1) Regulates the Contraction and Relaxation of Vascular Smooth Muscle and Maintains Blood Pressure

“…Blood Pressure Measurements-The systolic, diastolic, and mean blood pressures of conscious adult mice were recorded using a tail-cuff system (ALC-Non-invasive Blood Pressure System, Shanghai Alcott Biotech, China) as previously reported (13). Mice were trained by repeated blood measurement for 7 days before measurements were recorded.…”

“…Myography Assay for Vessel Contractility-The secondary branch of the mesenteric artery was isolated from adventitial tissues and mounted for isometric tension measurements (610-M, Danish Myo Technology) as previously reported (13). After equilibrating in pre-oxygenated HEPES-Tyrode buffer (137 mM NaCl, 2.7 mM KCl, 1.8 mM CaCl 2 , 1 mM MgCl 2 , 5.6 mM glucose, and 10 mM HEPES, pH 7.4) at 37°C for 30 min, the resting tension was adjusted to a value comparable to 100 mm Hg pressure in vivo.…”

“…Western blotting analysis was performed with a monoclonal antibody against myosin light chain (29). The percentage of phosphorylated myosin light chain relative to the total amount of myosin light chain was calculated as described previously (13).…”

“…3C). To assess arterial compliance in the MYPT1 SMKO mice, we compared the circumference corresponding to a transmural pressure of 100-mm Hg using a previously described method (13). IC 100 values for mesenteric arteries (n ϭ 9) were comparable between the MYPT1 SMKO and CTR vessels (362 Ϯ 20 m versus 361 Ϯ 14 m, p Ͼ 0.05).…”

“…Depolarization of the cell membrane by external stimuli activates voltage-gated Ca 2ϩ channels, resulting in Ca 2ϩ influx (5,6). Vascular tone is regulated via G-protein-coupled receptors (GPCRs) acting on two major signaling modules involving the heterotrimeric G proteins G q /G 11 and G 12 /G 13 . G q /G 11 mediates the activation of phospholipase C to generate inositol 1,4,5-trisphosphate, which releases Ca 2ϩ from the sarcoplasmic reticulum, leading to Ca 2ϩ /calmodulin-dependent activation of myosin light chain kinase (7)(8)(9).…”