Proceedings of The Physiological Society

Europhysiology 2018 (London, UK) (2018) Proc Physiol Soc 41, PCA355

Poster Communications

Fasudil modulates hypoxia-induced contraction in the isolated bovine digital vein: preliminary results

H. Zerpa1,2, A. Risso3,2, S. Comerma-Steffensen4,2

1. Department of Anatomy, Physiology and Pharmacology, School of Veterinary Medicine, St. George's University, St. George's, Grenada. 2. Department of Biomedical Sciences, Faculty of Veterinary Science, Central University of Venezuela, Maracay, Aragua, Venezuela, Bolivarian Republic of. 3. Department of Animal and Public Health, Faculty of Veterinary Medicine, National Experimental University "Romulo Gallegos", Zaraza, Guarico, Venezuela, Bolivarian Republic of. 4. Department of Biomedicine, Pulmonary and Cardiovascular Pharmacology, Aarhus University, Aarhus, Denmark.

  • Figure 1. Effect of fasudil (1μM-10μM) on the hypoxia-induced contraction in phenylephrine-contracted bovine digital veins. Mean ± S.E.M; n = 6. * p<0.05 significant difference between hyperoxia versus hypoxia; and † p<0.05 significant difference between fasudil 10μM versus hypoxia.

Hypoxia-induced contraction in the pulmonary vasculature is well described and calcium-sensitization, mediated by Rho kinase, is know to be a contributing factor (1). However, the effect of hypoxia in superficial veins has not been fully investigated. The bovine digital veins (BDVs) are superficial veins located in the subcutaneous tissue and drain blood from the claw. Isolated BDVs display hypoxia-induced contraction that is resistant to endothelium removal and extracellular calcium concentration depletion (2). The aim of the current study was to determine the effect of fasudil, a selective Rho kinase inhibitor, on isolated BDVs submitted to hypoxic conditions. BDVs were obtained from six cattle (Bos indicus) at a local abattoir and submitted to in vitro isometric contraction studies as previously described (2-3). Four parallel BDVs were used from the same animal and contracted to a similar degree with phenylephrine (0.6-1.0 µM) under hyperoxic conditions (95% O2- 5% CO2). After a submaximal contraction was achieved, four experimental groups were established: group 1-BDVs received no further treatment; group 2-BDVs were exposed to hypoxic conditions (95% N2 - 5% CO2) for 15 min; groups 3-BDVs & 4-BDVs were also exposed to hypoxic conditions but before adding phenylephrine, they were incubated with 1 µM fasudil and 10 µM fasudil respectively for 30 min. Responses were registered as: contraction (g/mg), percentage (%) of change over a basal submaximal contraction and area under the curve (AUC: arbitrary units). Negative and positive values outline isometric force below and above the basal contraction respectively. Values are presented as means ± S.E.M or 95% C.L (lower - upper), compared either by one-way or two-way ANOVA. The initial phenylephrine-induced contraction (g/mg) was similar (p>0.05) in all groups (group-1: 0.56 ± 0.1; group-2: 0.60 ± 0.1; group-3: 0.51 ± 0.03; group-4: 0.43 ± 0.07). Hypoxia caused an enhanced contractility (p<0.05) characterized by a peak contraction above the basal level at 5 min (Figure 1; group-1: 0.10% (-9.00 - 40.0); group-2: 24.0% (2.60 - 40.0); group-3: 20.7% (13.3 - 60.0); group-4: 5.01% (-23.0 - 64.5)). The total contractile response as AUC confirmed a significant (p<0.05) concentration dependent effect of fasudil on the hypoxia-induced contraction (group-1: -4387 (-13430 - 4657); group-2: 1164 (-6667 - 8994); group-3: 550 (-7770 - 8870); group-4: -8002 (-16248 - 243.8)). Hypoxia-induced contraction in BDVs was modulated by fasudil, Therefore, the Rho-kinase pathway could contribute to the observed hypoxia-induced contraction as was suggested in other veins under in vitro stressful conditions, such as reduction of temperature (3). The potential use of fasudil in clinical scenarios related to hypoxia in superficial veins of the limbs remains to be established.

Where applicable, experiments conform with Society ethical requirements