Tunicate Blood Cells

In the test for oxygen binding, 02-electrode measurements were made by diluting samples of oxygenated sea water with equal-sized volumes of either deaerated sea water or deaerated vanadocyte suspensions. If vanadocytes could bind 02, the concentration of dissolved 02 should be lower in the sample diluted with vanadocytes, than in the one diluted by deaerated sea water. However, no difference in the post-dilution sea water oxygen concentration was found between the two the results fell on the calibration curve. This finding has now been extended to tunicate blood cells rich in iron, rather than vanadium64. ... 

By using radioisotopes, the influx of vanadate into the blood cells of A. nigra has been studied85. The influx of phosphate, sulfate and dichromate, and the inhibitory effect of these oxoanions on vanadate influx have also been determined. The rate of vanadate influx was measured in the presence of metabolic inhibitors and inhibitors of anion transport. In this study, EPR spectroscopy was used to follow changes in the concentration of reduced vanadium within tunicate blood cells exposed to vanadate. 

C. Biological Origin Vanadium in Tunicate Blood Cells... 

Identification of different blood cell types is normally carried out using light (optical) microscopy. Blood cells are collected and are either fixed or spread alive on glass slides and then examined microscopically. Many types of blood cells have been identified, and relative numbers and types of cells have been found to differ among tunicate species. Tunicate blood cells are fragile and readily change appearance. It is possible that some commonly accepted blood cell types may actually be artifactual. 

Classification schemes have been devised that take into account the lability of tunicate blood cells (137,138). In one scheme (138) four main categories of tunicate blood cell are recognized stem cells, amoebocytes, vacuolated cells, and pigment cells (Fig. 4). Main categories are further subdivided in Ciona intestinalis, for example, hyaline amoebocytes are numerous and comprise 30% of the total blood cell population (139). 

The EPR technique has been used extensively to identify and characterize oxo-vanadium(IV) (i.e., vanadyl) in tunicate blood cells. Although EPR spectra are clearly diagnostic of vanadyl, different species yield different values of vanadyl EPR parameters (A and g values). Different values of EPR parameters have been reported for species such as A. ceratodes 144, 145), Leptoclinides lissus and Phallusia julinea 143). Vanadyl EPR Aq values of the blood cells of A. ahodori collected from different locations in the waters off Japan have been reported to differ from one another by approximately 10% (1.06 x 10 cm vs. 0.95 X 10 cm ), an amount about as much as interspecies differences 146). This result has been questioned, however, in the context of an extensive study of both Aplousobranch and Phlebobranch species utilizing several different methods to detect intracellular vanadium 147). 

Elements other than vanadium are accessible to the EXAS technique. Thus, abundant sulfur detected in tunicate blood cells has been identified as sulfate and sulfonate with EXAS (150). This result confirmed a similar finding carried out with chromatographic techniques (151). With this background, let us now consider the X-ray microprobe analyses of timicate blood cells. 

The occurrence of vanadium in the lower oxidation states, which as the simple aqua ions undergo acid dissociation above pH 3 [if present as V(III)) ] and pH 6 [in the case of oxo-V(IV) ], along with the high sulfur content of ascidian blood and the low pH that results when ascidian blood cells are ruptured in distilled water has led to the belief that intact vanadium-containing tunicate blood cells are acidic (145). Other lines of evidence, including vital staining and NMR (144,170), and... 

The issue of whether tunicate blood cells are involved in oxygen uptake and transport could, of course, be settled by experimentation. This challenge was met for tunicate specimens collected from both Atlantic " ] and Pacific ocean waters, " ] using an oxygen electrode to measure dissolved dioxygen concentrations. It was found that the oxygenbinding capacity of tunicate blood cells was indistinguishable from that of sea water. 

Agudelo, M.I., Kustin, K., McLeod, G.C., Robinson, W.E., and Wang, R.T. (1983) Iron accumulation in tunicate blood cells. I. Distribution and oxidation state of iron in the blood of Boltenia ovifera, Styda dava, and Molgula manhattensis. Biol. Bull., 165, 100-109. 

Dingley, A.L., Kustin, K., Macara, I.G., McLeod, G.C., and Robert, M.F. (1982) Vanadium-containing tunicate blood cells are not highly acidic. Biochim. Biophys. Acta, 720, 384-389. 

Kustin, K., Robinson, W.E., Erankel, R.B., and Spartalian, K. (1996) Magnetic properties of tunicate blood cells. II. Ascidia ceratodes. J. Inotg. Biochem., 63, 223-229. 

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