Differences in L-type Ca²⁺ channel activity partially underlie the regional dichotomy in pumping behavior by murine peripheral and visceral lymphatic vessels

Abstract

We identified a regional dichotomy in murine lymphatic contractile function with regard to vessel location within the periphery or visceral cavity. All vessels isolated from peripheral regions [cervical, popliteal, inguinal, axillary, and internodal inguinal axillary (Ing-Ax)] developed robust contractions with maximal ejection fractions (EFs) of 50-80% in our ex vivo isobaric myograph experiments. Conversely, vessels isolated from the visceral cavity (mesenteric, thoracic duct, and iliac) demonstrated maximal EFs of ≤10%. Using pressure myography, sharp electrode membrane potential recordings, and Ca imaging, we assessed the role of L-type Ca channels in this contractile dichotomy. Ing-Ax membrane potential revealed a ~2-s action potential (AP) cycle (resting -35 mV, spike -5 mV, and plateau -11 mV) with a plateau phase that was significantly lengthened by the L-type Ca channel agonist Bay K8644 (BayK; 100 nM). APs recorded from mesenteric vessels, however, displayed a slower upstroke and an elongated time over threshold. BayK (100 nM) increased the mesenteric AP upstroke velocity and plateau duration but also significantly hyperpolarized the vessel. Contractions of vessels from both regions were preceded by Ca flashes, detected with a smooth muscle-specific endogenous Ca reporter, that typically were coordinated over the length of the vessel. Similar to the membrane potential recordings, Ca flashes in mesenteric vessels were weaker and had a slower rise time but were longer lasting than those in Ing-Ax vessels. BayK (100 nM) significantly increased the Ca transient amplitude and duration in both vessels and decreased time to peak Ca in mesenteric vessels. However, a higher concentration (1 μM) of BayK was required to produce even a modest increase in EF in visceral lymphatics, which remained at <20%. NEW & NOTEWORTHY Lymphatic collecting vessels isolated from murine peripheral tissues, but not from the visceral cavities, display robust contractile behavior similar to lymphatic vessels from other animal models and humans. These differences are partially explained by L-type Ca channel activity as revealed by the first measurements of murine lymphatic action potentials and contraction-associated Ca transients.

Lymphatic Vessel Network Structure and Physiology

“…This includes the extreme case in most strains of mice, in which phasic contractions are so weak as to appear absent. Notably, the regional differences in the strength of lymphatic contractions in mice is related to differences in the activity of L-type Ca 2+ channels (1202). …”

T-type, but not L-type, voltage-gated calcium channels are dispensable for lymphatic pacemaking and spontaneous contractions

“…A segment of an excised lymphatic vessel containing at least one valve was transferred to a 3-mL chamber where it was cannulated onto two micropipettes and pressurized to 3 cmH 2 O. The bath solution containing Krebs buffer was exchanged at a rate of 0.5 mL/min and equilibrated for 30-60 minutes, at 37 °C, as previously described 52 . The inner diameter was tracked continuously from video images using a digital edge-detection method 54 .…”

Simplified method to quantify valve back‐leak uncovers severe mesenteric lymphatic valve dysfunction in mice deficient in connexins 43 and 37

“…Mesenteric lymphatic vessels were isolated as previously described Scallan et al 2016;Zawieja et al 2017;Castorena-Gonzalez et al 2018a,b;Zawieja et al 2018). Each vessel was pinned down with short segments of 40 µm stainless steel wire onto the Sylgard-coated surface of a dissection chamber filled with BSA containing Krebs buffer at room temperature.…”