Vanadocytes

Figure 17.16 Model of the pathway for reduction and accumulation of vanadium in ascidian vanadocytes. (From Crans et al., 2004. Copyright (2004) American Chemical Society.)...

The sea squirts or tunicates are fascinating marine creatures, their name being derived from the tunic made of cellulosic material that surrounds the body of the animal. In 1911, Henze discovered vanadium in the blood of Phallusia mammillata C.343 He later found the same with other ascidians (a class of tunicates). In vanadium-accumulating species, most vanadium is located in the vacuoles—vanadophores—of certain types of blood cells—the vanadocytes. The concentration in the vanadophore can be as high as 1M and this value must be compared with concentrations of the order of 2 x 10-8 M for vanadium in sea water.344 Kustin et al. have reviewed the work done to understand the efficient accumulation and the possible biological roles of the metal.345... 

For the vanadophore region, the sulfate concentration has been estimated to be of the order of 1.3 M and pH 2.349 With no evidence for a stable complex within the vanadocyte, Figure 14 depicts an interesting mechanism proposed for vanadium and sulfate accumulation.350 Anionic vanadium(V) (as HVO ") and sulfate ions enter the cell. Provided that within the vanadophore there is a strong reducing agent, vanadium can be reduced to vanadium(IV) and vanadium(III), cationic at the low pH in the vanadophore. If the vanadophore membrane is permeable to anions but not cations, the reduced vanadium remains trapped. 

The first suggestion of a possible biochemical function for vanadium came from the discovery that vanadocytes, the green blood cells of tunicates (sea squirts), contain 1.0 M V(III) and 1.5-2 M H2S04. 613 It was proposed that a V-containing protein is an oxygen carrier. However, the V3+ appears not to be associated with proteins612 and it does not carry 02. 

The ascidians or tunicates (sea squirts) accumulate vanadium from seawater (about 5x 10-8 mol dm-3) to a level of about 1 mol dm-3 and store it in a dilute solution of sulfuric acid (pH<2) in blood cells called vanadocytes. The tunicates thus concentrate vanadium several million-fold. 079 NMR, ESR and EXAFS determinations on whole vanadocyte cells of Ascidia ceratodes and Ascidia nigra indicate that the vanadium is present mainly as aquated V111 probably complexed with sulfate. Some vanadyl ion (5-10%) is also present.1080 1081... 

It was once thought that the vanadium in vanadocytes acted as dioxygen carriers but this is now known to be incorrect. Many questions remain to be answered about the role of vanadium in vanadocytes. 

Ueki, T., N. Yamaguchi, and H. Michibata. 2003. Chloride channel in vanadocytes of a vanadium-rich ascidian Ascidia sydneiensis samea. Comp. Biochem. Physiol. B Biochem. Mol. Biolog. 136 91-98. 

FIGURE 10.1 Ascidian vanadocyte. Modified from Reference 26. 

Yamaguchi, N., K. Kamino, T. Ueki, and H. Michibata. 2004. Expressed sequence tag analysis of vanadocytes in a vanadium-rich ascidian, Ascidia sydneiensis samea. Marine Biotech. 6 165-174. 

The chemistry of vanadium is characterized by multiple oxidation states (Fig. 1). The redox chemistry of this metal undoubtedly plays a role in its biochemistry. Of the four common oxidation states, only V(III), V(IV), and V(V) are important biologically, V(II) being too reducing to exist in any known organism. The best known example of the occurrence of V(III) is in the vanadocytes of the blood of tunicates22 otherwise, vanadium is largely found in the +4 and +5 oxidation states, both of which are readily accessible under physiological conditions. 

Extended X-ray absorption fine structure (EXAFS) and K edge absorption spectroscopy have been used to study vanadium in coaI7). In contrast to the common belief that vanadyl porphyrins are the major compounds present, these studies demonstrate that in the high vanadium content samples examined, the metal is tetravalent and coordinated to oxygen ligands. Similar investigations have been carried out on the vanadocytes of aci-dians to identify the form of vanadium in tunicate blood cells22 (vide infra). 

Only the features of vanadium biochemistry relevant to understanding the accumulation mechanism in vanadocytes are presented here. More extensive coverage of this subject may be found in two recent reviews51 525. 

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. ... 

Two types of evidence indicate that vanadium may function as a anti-feedant for unicates. First, there is evidence based on observation that species whose outer coverings (tunics) are rich in vanadocytes are shunned by predatory fish65,66. These tunics are often acidic, which confers further protection from predation on an individual specimen exhibiting this characteristic. Since vanadium would be expected to hydrolyze in sea water, this acidity could result from release of vanadium due to rupturing blood cells. Second, pieces of fish deliberately dosed with appreciable amounts of vanadium, and offered as food pellets to crustaceans and fish, resulted in reduced food consumption by these predators67. ... 

This paper reported cellular responses in Molgula manhattensis, a species which contains ferro-cytes, but no vanadocytes. We have repeated the experiments with C. intestinalis, a vanadocyte-containing species, and found responses similar to those described for M. manhattensis... 

Fig. 2. Electron micrograph of vanadocytes from Ascidia nigra. Cells were fixed in 2.5% glutaral-dehyde in 0.15 M HC1, followed by 1% osmium tetroxide, pH 7.6. The sections were stained with uranyl acetate N = nucleus, PM = plasmalemma, V = vacuoles (very electron dense presumably because of reduction of osmium tetroxide by V(III).)...

Vanadocytes are nucleated, possess organelles, although few mitochondria80 and the conspicuous vacuoles. The cells of many species such as A. nigra8>> and A. ceratodes82> are a bright yellow-green, due to the vacuolar coloration. These observations are drawn from microscopic examination of blood samples. It is more difficult to isolate vanadocytes from all other blood cells, and determine their chemical composition. However,... 

To avoid these pitfalls two different techniques have recently been used to measure the intracellular pH in intact A. nigra blood cells97. One technique combines a measurement of the ratio of intracellular to extracellular concentration of amine with measurement of the extracellular pH to yield the intracellular pH. Another technique involves intracellular 31P-NMR (nuclear magnetic resonance) measurements. These methods led to a determination of pH7.39 0.10 for unfractionated cells, and 7.2 for the vanadocyte fraction. 

The influx of vanadate is a rapid (t1/2 = 57 s at 0°C), monophasic process85, but the rate of vanadate influx approaches a saturation limit at high concentrations of exogenous vanadate. Influx experiments with added EDTA (ethylenediamminetetraacetic acid) were carried out to eliminate exchange or adhesion of vanadate to the cell surface to account for the reduction of radiovanadium in the extracellular medium. The amount of vanadium accumulated in a typical influx experiment was independent of added EDTA, and raised the intracellular concentration by 2.5 mM. Therefore, there is a net uptake of vanadium into the intracellular volume of the vanadocytes in these experiments. 

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