Hypoxia-induced pulmonary arterial contraction appears to be dependent on myosin light chain phosphorylation
Zhao, Y.; Rhoades, R.A.; Packer, C.S.
American Journal of Physiology 271(5 Pt 1): L768-L774
1996
ISSN/ISBN: 0002-9513 PMID: 8944720 Document Number: 467770
The signal transduction pathway of hypoxic pulmonary arterial contraction has not been elucidated. Phosphorylation of the 20-kDa myosin light chain (MLC-20) is thought to be essential for vascular muscle contraction. However, there are reports that smooth muscle will contract in response to nonphysiological stimuli such as phorbol esters without the involvement of MLC-20 phosphorylation. The purpose of this study was to determine if hypoxia-induced pulmonary arterial contraction is dependent on MLC-20 phosphorylation. Isolated rat pulmonary and carotid (for comparative purposes) arterial strips were contracted with 80 mM KCl to establish maximum active tension in response to membrane depolarization. The strips were then stimulated with one of the following: 30 mM KCl, 1 mu-M phenylephrine, 0.01 mu-M angiotensin II, 1 mu-M phorbol 12-myristate 13-acetate (PMA), or hypoxia (95% N-2-5% CO-2). In some experiments ML-9, a myosin light chain kinase inhibitor, or calphostin C, a protein kinase C (PKC) inhibitor, was introduced into the bath before hypoxia. Isometric tension was recorded as a function of time. Muscle strips were freeze-clamped (liquid N-2) at various time points during the course of responses to the various stimuli. MLC-20 phosphorylation levels were measured by urea-glycerol gel electrophoresis followed by Western blot procedure. Results show that increased MLC-20 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. The MLC-20 phosphorylation levels correlate with tension development in response to hypoxia, and ML-9 abolished the hypoxic contractions. In contrast, hypoxia relaxed carotid arterial muscle, and there was a corresponding decrease in the MLC-20 phosphorylation level. In conclusion, hypoxia appears to result in MLC-20 phosphorylation-mediated contraction in conduit pulmonary arterial muscle and in MLC-20 dephosphorylation-mediated relaxation in systemic arterial muscle.