Lymphatic muscle contraction is critical for the centripetal movement of lymph that regulates fluid balance, protein homeostasis, lipid absorption and immune function. We have demonstrated that lymphatic muscle has both smooth and striated muscle contractile elements; however, the basic contractile properties of this tissue remain poorly defined. We hypothesized that contractile characteristics of lymphatic myofilaments would be different from vascular smooth muscle myofilaments. To test this hypothesis, pCa-tension relationship was determined for -toxin permeabilized mesenteric lymphatics, arteries and veins. The Ca²⁺ sensitivity (pCa₅₀) of lymphatics was significantly lower compared to arteries (6.16±0.05 vs. 6.44±0.02; p<0.05), whereas there was no difference in pCa₅₀ between lymphatics and veins (6.16±0.05 vs. 6.00±0.10; n.s.). The Hill coefficient for -toxin permeabilized lymphatics was not significantly different from arteries, but was significantly greater than that of the veins (1.98±0.19 vs. 1.21±0.18; p<0.05). In addition the maximal tension and pCa₅₀ values were significantly greater in -toxin permeabilized lymphatics compared to -escin permeabilized lymphatics (0.27±0.03 vs. 0.15±0.01 mN/mm and 6.16±0.05 vs. 5.86±0.06, respectively; p<0.05), whereas the Hill coefficient was significantly greater in -escin permeabilized lymphatics. Western blot analyses revealed that CPI-17 levels were significantly decreased by about 50% in -escin permeabilized lymphatics, compared to controls, whereas no change in the level of calmodulin was detected. Our data constitute the first description of the pCa-tension relationship in lymphatic muscle. It suggests that differences in myofilament Ca²⁺ sensitivity and cooperativity among lymphatic muscle and vascular smooth muscles contribute to the functional differences that exist between these tissues.