Hypoxia-induced pulmonary arterial contraction appears to be
dependent on myosin light chain phosphorylation.
Zhao, Ying, Rodney A. Rhoades, and C. Subah Packer.
Department of Physiology and Biophysics, Indiana University School
of Medicine, 635 Barnhill Drive, Indianapolis, Indiana 46202-5120
APStracts 3:0108L, 1996.
Hypoxic pulmonary arterial contraction is a direct response of
pulmonary arterial muscle to hypoxia but the signal transduction
pathway has not been elucidated. Phosphorylation of the 20 kD myosin
light chain (MLC20) is thought to be essential for vascular muscle
contraction in response to most physiological agonists. However,
there are reports that smooth muscle will contract in response to
other stimuli such as phorbol esters without the involvement of MLC20
phosphorylation. The purpose of this study was to determine if
hypoxia-induced pulmonary arterial contraction is dependent on MLC20
phosphorylation. Carotid arterial muscle was investigated for
comparative purposes. Isolated rat pulmonary and carotid arterial
strips were equilibrated in tissue baths and contracted with 80mM KCl
to establish maximum active tension in response to membrane
depolarization (P0). The strips were then stimulated with one of the
following: 30mM KCl, 1uM phenylephrine, 0.01uM angiotensin II, 1uM
phorbol myristic acetate (PMA) or hypoxia (95%N2/5% CO2). In some
experiments ML-9, a myosin light chain kinase (MLCK) inhibitor, was
introduced into the bath prior to hypoxia. In some other cases,
calphostin C, a protein kinase C (PKC) inhibitor, was utilized.
Isometric tension was recorded as a function of time. Muscle strips
were freeze-clamped (liquid N2) at various time points during the
course of responses to agonist stimulation or hypoxia. MLC20
phosphorylation levels were measured by urea-glycerol gel
electrophoresis followed by Western blot procedure. Results show that
increased MLC20 phosphorylation correlates with initiation of
pulmonary arterial smooth muscle contraction in response to all
agonists with the exception of PMA, a known activator of PKC. PMA
induces an increase in tension without a change in MLC20
phosphorylation levels. The MLC20 phosphorylation levels correlate
with tension development in response to hypoxia and ML-9 abolished
the hypoxic contractions while calphostin C had varying effects on
hypoxia phase 1 and 2 contractions. In contrast, hypoxia relaxed
carotid arterial muscle and there was a corresponding decrease in
MLC20 phosphorylation level. In conclusion, hypoxia appears to result
in MLC20 phosphorylation- mediated contraction in conduit pulmonary
arterial muscle and in MLC20 dephosphorylation-mediated relaxation in
systemic arterial muscle.
Received 20 January 1995; accepted in final form 20 June 1996.
APS Manuscript Number L15-5.
Article publication pending Am. J. Physiol. (Lung Cell. Mol.
Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 4 July 96