Davis MJ, Lane MM, Davis AM, Durtschi D, Zawieja DC, Muthuchamy M, Gashev AA. Modulation of lymphatic muscle contractility by the neuropeptide substance P. Am J Physiol Heart Circ Physiol 295: H587-H597, 2008. First published June 6, 2008 doi:10.1152/ajpheart.01029.2007.-Substance P (SP) is a neuropeptide associated with sensory innervation of lymphoid tissue and a suspected modulator of lymphatic function in inflammation. Only a few studies have examined the effects of SP on lymphatic contraction, and it is not clear to what extent SP acts directly on the lymphatic muscle and/or endothelium or indirectly through changes in intraluminal filling pressure secondary to increases in capillary permeability/ filtration. We tested the effects of SP on the spontaneous contractions of rat isolated mesenteric lymphatic vessels under isometric and isobaric conditions, hypothesizing that low concentrations would stimulate lymphatic pumping by enhancing lymphatic muscle contraction in a manner complementary to the effect of increased preload. Under isometric conditions, SP (10 nM) dramatically enhanced lymphatic chronotropy and inotropy. Unlike guinea pig lymphatics, SP actions were not blocked by cyclooxygenase or PLA2 inhibition. In the absence of SP, ramp increases in isometric preload resulted in ϫϳ1.6 increases in contraction amplitude (Amp) and ϫϳ1.7 increases in frequency (Freq). SP increased Freq by ϫϳ2.4, Amp by ϫϳ1.9, and the Amp-Freq product (AFP) by ϫϳ3.5. Under isobaric conditions, the pressure elevation from 0.5 to 10 cmH2O in the absence of SP decreased Amp by ϫϳ0.6 and increased Freq by ϫϳ1.8. SP caused a modest increase in Amp, a robust increase in Freq at all pressures, and shifted the AFP-pressure relationship upward and leftward. Therefore, SP has substantial positive inotropic and chronotropic effects on rat lymphatic muscle, improving pump efficiency independent of the effects of preload and broadening of the working range of the lymphatic pump. neurokinin; edema; inflammation; lymphatic pump; inotropy; chronotropy; thromboxane A2 SUBSTANCE P (SP) is an 11-amino acid neuropeptide (36) often associated with cells of lymphoid tissue (15,19,21,22,46,47). SP is released from enteric nerves, sensory nerves, and inflammatory cells (27,41,45,52). Correspondingly, SP receptors are expressed on endothelium and muscle of blood and lymphatic vessels, on nerves innervating those vessels, and on associated immune and inflammatory cells (31,36,44).SP appears to have both direct and indirect effects on vascular targets. For example, SP is a powerful stimulator of endothelium-dependent nitric oxide production (28), evoking vasodilation and increasing local blood flow. In addition, SP enhances vascular permeability (18,25) and stimulates the release of cytokines and chemokines (4, 36) that would indirectly elicit vasodilation and increase vascular permeability. These vascular effects of SP would collectively increase flow/ pressure in capillaries and post-capillary venules and lead to increases in transcapillary fluid ...

Isometric and isobaric methods yielded qualitatively similar indices of spontaneous contractile activity. However, the ranges of amplitude and frequency changes were much greater under isobaric conditions (3- to 5-fold) than under isometric conditions (50-80%).

Phasic contractile activity in rat portal vein is more sensitive to the rate of change in length than to absolute length and this response is widely assumed to be a general characteristic of myogenic behaviour for vascular smooth muscle. Previously, we found that rat lymphatic vessels exhibit phasic contractile behaviour similar to that of portal vein. In the present study, we hypothesized that lymphatic muscle would exhibit rate-sensitive contractile responses to stretch. The hypothesis was tested on rat mesenteric lymphatics (90-220 μm, i.d.) using servo-controlled wire-and pressure-myograph systems to enable ramp increases in force or pressure at different rates. Under isometric conditions in wire-myograph preparations, both the amplitude and the frequency of phasic activity were enhanced at more optimal preloads, but superimposed upon this effect were bursts of contractions that occurred only during fast preload ramps. In such cases, the ratio of contraction frequency during the ramp to that at the subsequent plateau (at optimal preload) was > 1. Further, the frequency ratio increased as a function of the preload ramp speed, consistent with a rate-sensitive mechanism. In contrast, the amplitude ratio was < 1 and declined further with higher ramp speeds. Downward preload ramps produced corresponding rate-sensitive inhibition of contraction frequency but not amplitude. Similar findings were obtained in pressurized lymphatics in response to pressure ramps and steps. Our results suggest that lymphatics are sensitive to the rate of change in preload/pressure in a way that is different from portal vein, possibly because the pacemaker for generating electrical activity is rate sensitive but lymphatic muscle is not. The behaviour may be widely present in collecting lymphatic vessels and is probably an important mechanism for rapid adaptation of the lymphatic pump to local vascular occlusion. Phasic contractile activity of smooth muscle in rat portal vein is known to be more sensitive to the rate of change in length than to the absolute length (Johansson & Mellander, 1975). Under isometric conditions, the longitudinal muscle layer of portal vein exhibited spontaneous contractile activity that was superimposed on, and modulated by, the level of passive force (preload). When preload was elevated gradually from suboptimal to optimal level, large amplitude (AMP), high frequency (FREQ) contraction bursts were evident in the force recordings (Johansson & Mellander, 1975). The most prominent contraction bursts occurred just before and just after passive force reached a plateau at a more optimal preload. During the plateau, both AMP and FREQ This paper has online supplemental material. declined somewhat to values that were still higher than their respective values at the original (suboptimal) preload. Thus, the values of both AMP and FREQ were larger during the force ramp than at the higher, plateau level associated with a more optimal preload. Portal vein also exhibited the opposite response to reduction in preload: during ...

The method works well with isometric rat mesenteric lymphatics without disturbing spontaneous activity. It should be applicable to arterial, venous, and lymphatic vessels (80-500 microm in diameter) isolated from other tissues and species.

Both the amplitude (AMP) and frequency (FREQ) of phasic contractile activity in rat portal vein are more sensitive to the rate of length change than to the absolute length (Circ Res 36:, 1975). Because lymphatics exhibit similar phasic activity, we hypothesized that lymphatic muscle would exhibit rate‐sensitive responses to stretch. Rat mesenteric lymphatics (~100 um, ID) were studied in vitro using customized wire‐ and pressure‐ myographs that allowed servo‐control of diastolic force, diameter, or pressure (preload). While recording spontaneous activity, preload was increased at variable rates, using preload ramps with different slopes, from the lowest value at which spontaneous activity occurred to the optimal level. Both contraction AMP and FREQ increased with preload, as reported previously. Superimposed upon this were bursts of contractions during the rising preload phase. The ratio of contraction FREQ (ramp/end) was always greater than 1 and increased with rate. In contrast, the AMP ratio was always less than 1 and was independent of rate. Off‐responses (contraction inhibition) were typically more pronounced than on‐responses. These results suggest that lymphatic vessels are sensitive to the rate of change of preload in a different way than portal vein. One explanation is that the electrical pacemaker in the lymphatic wall is rate‐sensitive but the muscle itself is not. Supported by NIH HL‐75199.