Inhibition of myosin light chain phosphorylation decreases rat mesenteric lymphatic contractile activity

Abstract

Muscular lymphatics use both phasic and tonic contractions to transport lymph for conducting their vital functions. The molecular mechanisms regulating lymphatic muscle contractions are not well understood. Based on the well-established finding that the phosphorylation of myosin light chain 20 (MLC20) plays an essential role in blood vessel smooth muscle contraction, we investigated if phosphorylated MLC20(pMLC20) would modulate the tonic and/or phasic contractions of lymphatic muscle. The effects of ML-7, a MLC kinase inhibitor (1–10 μM), were tested on the contractile parameters of isolated and cannulated rat mesenteric lymphatics during their responses to the known modulators, pressure (1–5 cmH2O) and substance P (SP; 10−7M). Immunohistochemical and Western blot analyses of pMLC20were also performed on isolated lymphatics. The results showed that 1) increasing pressure decreased both the lymphatic tonic contraction strength and pMLC20-to-MLC20ratio; 2) SP increased both the tonic contraction strength and phosphorylation of MLC20; 3) ML-7 decreased both the lymphatic tonic contraction strength and pMLC20-to-MLC20ratio; and 4) the increase in lymphatic phasic contraction frequency in response to increasing pressure was diminished by ML-7; however, the phasic contraction amplitude was not significantly altered by ML-7 either in the absence or presence of SP. These data provide the first evidence that tonic contraction strength and phasic contraction amplitude of the lymphatics can be differentially regulated, whereby the increase in MLC20phosphorylation produces an activation in the tonic contraction without significant changes in the phasic contraction amplitude. Thus, tonic contraction of rat mesenteric lymphatics appears to be MLC kinase dependent.