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Vascular control in larval Xenopus laevis: the role of endothelial-derived factors

Thorsten Schwerte^*, Eva Printz and Regina Fritsche

Department of Zoophysiology, University of Göteborg, Sweden

View larger version (87K): [in a new window]   Fig. 1. (A,B) Video images showing a cast of the vascular bed of the main vessels in the head of a Xenopus laevis tadpole using the digital motion-analysis technique. In the anaesthetized tadpole, the movements of the erythrocytes are digitally contrasted. The arrows point to the location where vessel diameter measurements were made by defining a threshold for the grayscale value and applying the ‘rectangle-fit’ algorithm. This automatically defined the best-fitting rectangle covering the blood vessel within this region of interest. The length of the short axis of this rectangle was taken as vessel diameter (see Materials and methods for details). (A) Before endothelin application; (B) after endothelin (ET-1, 10–6 mol l–1) application. ha, head artery. Image width, 1.6 mm.

View larger version (82K): [in a new window]   Fig. 2. (A–C) Endothelin-1-like immunoreactivity (ET-1 IR) in Xenopus laevis NF stage 50 tadpoles. Cross sections of the head region cranial to the heart and truncus arteriosus with the ventral side at the top of the figure. (A) The immunoreactive endothelial cells of the main head arteries (arrows). Note that the two main head veins lack ET-1 IR (arrowheads). The body wall showed nonspecific labelling (top), which was also present in control sections. Scale bar, 400 µm. (B) The two main head arteries with the endothelium, showing ET-1 IR. The smooth muscle layer, the connective tissue and the melanophores covering parts of the blood vessels cannot be seen in this micrograph. Scale bar, 200 µm. (C) A higher-power view of one of the main head arteries. Scale bar, 100 µm.

View larger version (26K): [in a new window]   Fig. 3. (A) Effects of endothelin (ET-1) on the diameter of the main artery to the head (normalized to the diameter under control conditions) in Xenopus laevis tadpoles (N-F stages 50–53). Values are means ± S.E.M. (N=6); the symbols and asterisks indicate significant differences from the control period. (B) Effect of endothelin (ET-1) on the diameter of the main vein from the head (normalized to the diameter under control conditions) in Xenopus laevis tadpoles. Values are means ± S.E.M. (N=6). Arrowheads indicate the point at which endothelin was applied.

View larger version (24K): [in a new window]   Fig. 4. Effects of 10–7 mol l–1 endothelin (ET-1) on the normalized diameter of the main artery to the head (normalized to the diameter under control conditions) in Xenopus laevis tadpoles. One group of tadpoles was preincubated for 20 min (black bar) with L-NAME (10–4 mol l–1) in Ringer’s solution prior to injection of endothelin. The other group was preincubated with Ringer’s solution alone for 20 min (black bar) prior to injection of endothelin. The arrowhead indicates the time of endothelin injection. Values are means ± S.E.M. (N=6); symbols and asterisks indicate significant differences from the control period.

View larger version (22K): [in a new window]   Fig. 5. Effects of 10–6 mol l–1 endothelin (ET-1) and 10–2 mol l–1 sodium nitroprusside (SNP) on the normalized diameter of the main artery to the head (normalized to the diameter under control conditions) in Xenopus laevis tadpole. Arrowheads indicate the time of drug injections. Values are means ± S.E.M. (N=6). Asterisks indicate significant differences from the control period.

View larger version (36K): [in a new window]   Fig. 6. (A) Schematic drawing showing the possible mechanism of regulation of vessel diameter of the main head artery in Xenopus laevis tadpoles before functional innervation of the peripheral vascular system. Endothelin (ET-1) is known to be a potent and long-lasting endothelium-derived vasoconstrictor. Nitric oxide (NO) can also be produced by the endothelium and has a dilatory effect. NO is known to be released in response to shear stress induction. Recent studies (see text, for references) provide evidence for a functional coordination and cooperation between NO and ET-1. The time course is indicated on the axis in B. The numbers in circles indicate the corresponding points in B. (1) Endothelin, possibly released by the endothelium, provokes a strong vasoconstriction, which in turn indirectly induces the release of NO (3) by increasing shear stress (2). NO acts directly on the vascular smooth muscle (vsm) to dilate the vessel (5). In addition, it acts as a ‘physiological brake’ on endothelin function by decreasing the affinity of ET-1 for its receptor (4), which enhances the vasodilation (5). L-NAME is a competitive inhibitor of the endothelial nitric oxide synthase. (6) The effect of preincubation with L-NAME. (B) Typical pharmacological experiments demonstrating the possible coordination and cooperation between NO and ET-1. SNP, sodium nitroprusside.