Control of lymphatic pacemaking and pumping by mechanobiological signals

The spontaneous, phasic contractions of collecting lymphatic vessels are critical for lymph transport and interstitial fluid homeostasis. Phasic contractions are initiated by action potentials in lymphatic muscle and conduct along the vessel to trigger contraction waves. Contractions are regulated by pressure and shear stress (SS), but only limited aspects of that regulation are understood. Numerical models predict that pressure promotes retrograde propagation of contraction waves, whereas nitric oxide (NO) production associated with phasic contractions (pulsatile NO) promotes antegrade conduction and extends the pressure range over which contractions propel lymph. These predictions were tested using 3-4-valve segments of rat mesenteric lymphatic vessels using pressure myography and protocols that imposed forward flow, elevated inflow pressure (P_in) or elevated outflow pressure (P_out), each with/without intact NO signalling. NO bioavailability and flow-induced responses were enhanced by l-arginine supplementation. Spatiotemporal maps generated from video images were used to quantify the direction and extent of contraction wave conduction. Our results show that (1) contraction waves are normally biased towards retrograde conduction at equal P_in/P_out levels. (2) P_in elevation promotes antegrade conduction, whereas P_out elevation promotes retrograde conduction. (3) Imposed flow is inhibitory, reducing contraction amplitude and frequency and limiting the extent of contraction wave conduction without a significant effect on conduction direction. (4) Pulsatile NO does not significantly influence the conduction direction or extend the pressure range over which spontaneous contractions occur. Our findings support the idea that pressure is the dominant regulator of lymphatic pacemaking and pumping, with pulsatile NO having only minimal influence. KEY POINTS: The degree to which spontaneous, phasic contractions of lymphatic collecting vessels are regulated by pressure and shear stress is not fully understood. Numeric models predict that nitric oxide (NO) production associated with phasic contractions (pulsatile NO) promotes antegrade conduction of contraction waves, whereas pressure elevation promotes retrograde conduction; pulsatile NO production is also thought to extend the pressure range over which phasic contractions occur. Ex vivo methods were used to control pressure/flow in 3-4 valve segments of collecting lymphatics from rat mesentery, with preserved or inhibited NO signalling. The relatively long vessel segments limited the absolute levels of imposed flow/SS, so l-arginine supplementation was used to enhance NO bioavailability. Our findings support a scheme whereby pressure is by far the dominant mechanism determining the pacemaking site of lymphatic collectors, and challenge existing dogma about the importance of pulsatile NO production in regulating their behaviour.