Concomitant increase in cytosolic free calcium and phosphorylation of myosin light chain by vasoconstrictive hormones in cultured rat vascular smooth muscle cells.
Using the fluorescent Ca2+ indicator quin-2 to measure the cytosolic Ca2+ concentration [( Ca2+]i) and glycerol-urea polyacrylamide gel electrophoresis combined with radioimmunoblotting to assess phosphorylation of 20 k-dalton (Da) myosin light chain (MLC), the effects of angiotensin (A) II and arginine-vasopressin (AVP), both potent vasoconstrictive hormones, on changes in [Ca2+]i and phosphorylation levels of 20 k-Da MLC were studied in cultured rat vascular smooth muscle cells (VSMC). AII and AVP induced immediate (within 1 min) and dose-dependent increases in both [Ca2+]i and phosphorylation of 20 k-Da MLC. Pretreatment of VSMC with AII receptor antagonist and V1 receptor antagonist, completely blocked increases in [Ca2+]i by AII and AVP, respectively. Phosphorylation of 20 k-Da MLC stimulated by these agonists was also inhibited by their specific antagonists. These data suggest that receptor-mediated increases in [Ca2+]i by AII and AVP are closely associated with phosphorylation of 20 k-Da MLC in VSMC. The present methods should provide a suitable in vitro cell model for investigation of the molecular mechanism by which vasoactive hormones act on cells to induce contraction of vascular smooth muscle.
Takata
S
S
Hypertension-Endocrine Division, National Cardiovascular Center Research Institute, Osaka, Japan.
Hirata
Y
Y
Furukawa
K
K
Tomita
M
M
Takagi
Y
Y
Fujita
T
T
Hidaka
H
H
D000634:Aminoquinolines; D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D001127:Arginine Vasopressin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism*; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism*; D010766:Phosphorylation; D011863:Radioimmunoassay / Q000379:methods; D051381:Rats; D011919:Rats, Inbred Strains; D014662:Vasoconstrictor Agents / Q000494:pharmacology*
D000634:Aminoquinolines; D014662:Vasoconstrictor Agents; D000804:Angiotensin II; D001127:Arginine Vasopressin; D009218:Myosins; C035739:Quin2; D002118:Calcium