The intracellular distribution of ions and water in rat liver and heart muscle

Local dry mass concentrations of intracellular compartments in rat heart muscle and liver cells were estimated by quantitative electron microscopy and X-ray microanalysis of ultrathin frozen-dried cryosections. The results were used to calculate elemental concentrations per litre of compartment water from the X-ray microanalytical data. Water fractions were between 80.3 ± 1.3% of wet weight in the decondensed chromatin and only 45.1 ± 1.7% in mitochondria of liver cells. The lowest water fraction in heart muscle cells was also found in mitochondria. The ionic concentrations found in the cytoplasm of liver cells and in the myofibrils are in accord with the electroneutrality rule and in osmotic equilibrium with the extracellular concentrations. The concentrations of Na, K, Cl and P both in the cytoplasm and in the regions of decondensed chromatin within the nuclei were found to be equal. However, in regions of condensed chromatin K⁺ concentrations were found to be much higher than expected for a Donnan distribution of ions free in solution. Most probably the activity coefficient for K⁺ is lower in the condensed chromatin than in the decondensed or in the cytoplasm. The same holds true for the A-band as compared to the I-band in heart muscle cells. A sequestration of K⁺ was measured also in the rough endoplasmic reticulum (RER) of hepatocytes. The Cl^? concentration in mitochondria both in heart muscle and liver cells has been measured far in excess of what might be expected from a Nernstian distribution. A coupled inward Cl^? transport in mitochondria must, therefore, be assumed.     

Gender-related changes in the avian vasotocin system during ontogeny

The arginine vasotocin (AVT) system of the avian brain includes a sexually dimorphic part that extends from the caudal part of preoptic region through the medial part of the bed nucleus of stria terminalis (BSTm) to the lateral septum. It is composed of the parvocellular neurons located in the BSTm and the dense innervation of the medial preoptic region and lateral septum. In this part of the brain, AVT expression is stronger in males than in females in a few bird species investigated to date. This review focuses on the ontogeny of sexual differences in the vasotocinergic system of two gallinaceous species, domestic chicken and Japanese quail, and on the role of gonadal hormones in organizing during development and maintaining in adulthood these differences. Parvocellular AVT neurons become discernible in the BSTm of males and females during the second half of embryonic development. These cells undergo a profound and irreversible sexual differentiation during ontogenetic development. Recent findings demonstrate a dual role of estrogens in the organization and activation of sex differences in the AVT system. During the embryonic period of ontogeny, estrogens differentiate the AVT system in a sexually dimorphic manner in parallel with the differentiation of sexual behavior, while in adulthood estrogens, locally produced from testosterone in the male brain, activate AVT synthesis in the BSTm. The sexually dimorphic part of the AVT system is sensitive to a number of abiotic factors such as light, temperature, and water availability. It is suggested that sex dimorphic vasotocinergic systems could be implicated in processes of social recognition in various behavioral contexts.     

Ciliary neurotrophic factor increases the survival of magnocellular vasopressin and oxytocin neurons in rat supraoptic nucleus in organotypic cultures.

Organotypic cultures of the rat hypothalamus are very useful models for the long-term study of parvocellular vasopressin (VP) neurons in the paraventricular (PVN) and suprachiasmatic (SCN) nuclei. However, they do not preserve significant numbers of VP magnocellular neurons (VP-MCNs) in either the PVN or the supraoptic nucleus (SON). Vutskits et al. [(1998) Neuroscience 87:571-582] reported that ciliary neurotrophic factor (CNTF) was a selective survival factor for rat VP-MCNs in organotypic cultures of the rat hypothalamic paraventricular nucleus (PVN). We examined the effects of CNTF on the survival of these neurons in rat and mouse SONs. CNTF (10 ng/ml) in the culture media increased the survival of VP-MCNs by 6-fold and OT-MCNs by 3-fold. In the mouse, both OT- and VP-MCNs survive very well in organotypic cultures under standard culture conditions and the addition of CNTF had no further effect. Consistent with these results, in situ hybridization showed substantially higher levels of VP- and OT-mRNA in rat PVNs and SONs in the presence of CNTF, but produced no changes in these nuclei in the mouse. The optimum period for the survival effect of CNTF on MCNs in the rat hypothalamic cultures was in the first 7-10 days of culture and this effect is maintained for at least 5 additional days if CNTF is then removed from the medium. Therefore, using CNTF in the culture media can provide an opportunity for long-term studies of rat VP- and OT-MCNs in SONs in organotypic cultures.     

Sex differences in songbirds 25 years later: What have we learned and where do we go?

About 25 years ago, Nottebohm and Arnold reported that there are profound male-biased sex differences in volume in selected nuclei in telencephalic portions of the song control system. This review focuses on issues related to the cellular bases of these sex differences in volume and comparative studies that might elucidate the function of this variation between the sexes. Studies utilizing a variety of neurohistological methods in several different species to define the boundaries of two key telencephalic song nuclei HVc and the robust nucleus of the archistriatum (RA) all tend to find a sex difference in volume in agreement with Nissl-defined boundaries. Sex differences in volume in nuclei such as HVc and RA are associated with differences in cell size and cell number. Other attributes of the phenotype of cells in these nuclei are also different in males and females such as the number of cells expressing androgen receptors. Comparative studies have been employed to understand the function of these sex differences in the brain. In some songbird species, females sing rarely or not at all, and the brain nuclei that control song are many times larger volume in males than females. In other species, males and females sing approximately equally, and the brain nuclei that control song are approximately equal between the sexes. Recently, statistical methods have been employed to control for phylogenetic effects while comparing the co-evolution of traits. This analysis indicates that the evolution of sex differences in song has co-evolved with the evolution of sex differences in singing behavior in songbird species. Future studies should focus on the function of the smaller song control nuclei of females and investigate the role these nuclei might play in perception as well as in production. Microsc. Res. Tech. 54:327–334, 2001. © 2001 Wiley-Liss, Inc.     

Dynamic behaviors of nonlinear fractional-order differential oscillator

The nonlinear dynamic behaviors of oscillators described by fractional-order differential are presented in this paper. The background of the research is based upon two engineering practices. First is that the visco-elastic behaviors of some advanced polymeric materials can be accurately modeled by the fractional calculus constitutive law. Second is that the influence of nonlinear visco-elasticity described by the fractional operator cannot be neglected in some cases such as the vibration with large displacement or large strain and thermo-visco-elastic coupled problems. The numerical scheme for solving the nonlinear equation of motion is developed. The results show that because of the introduction of nonlinear damping modeled by the fractional-order operator, the bifurcation and chaos of the oscillator appear in forced vibration. Furthermore, the fraction value of the fractional operator evidently affects the dynamic behavior of the nonlinear fractional differential oscillator. This paper was presented at the 4th Asian Conference on Multibody Dynamics(ACMD2008), Jeju, Korea, August 20–23, 2008. Wei Zhang received his doctorate degree in Mechanics at Xi’an Jiaotong University in China in 1994. He went to the University Of Sheffield in the UK as an academic visitor from July 1986 to July 1987. He completed his post-doctorate study in Iwaki Meisei University in Japan through the support of the JSPS from May 1997 to June 1999. Dr. Zhang is currently a professor at the Department of Electronic Engineering of JiNan University in Guangzhou, China. Shao-kai Liao received his B.E. degree in Civil Engineering at JiNan University in Guangzhou, China in 2004 and his master’s degree in Engineering Mechanics at the same university in 2006. Mr. Liao was once a postgraduate student of Professor Wei Zhang, and he now works in an institute in Shenzhen City in China. Nobuyuki Shimizu received his B.S. degree in Mechanical Engineering at Yokohama National University and his master’s and doctorate degrees in Mechanical Engineering at University of Tokyo in 1968 and 1971, respectively. He went to the California Institute of Technology in California, U.S.A. as a visiting associate from October 1976 to September 1977. Dr. Shimizu is currently a professor at the Department of Mechanical and Power Engineering of Iwaki Meisei University in Japan.     

A synchrotron X-ray study of the changes occurring in the corneal stroma during processing for electron microscopy

Using a high-intensity synchrotron X-ray source, the structural changes occurring in the corneal stroma were monitored during each stage of several different processing runs for the transmission electron microscope (TEM) and scanning electron microscope (SEM). The parameters studied were interfibrillar spacing, intermolecular spacing, D-periodicity and fibril diameter. The processing schedule that produced the least changes in spacings for TEM specimens involved extended fixation in glutaraldehyde followed by low-temperature embedding in Lowicryl K4M resin. However, interfibrillar material was better preserved after embedding in LR White resin or Nanoplast. Almost every processing stage for electron microscopy produced significant changes in one or more structural parameters in the cornea. Glutaraldehyde fixation significantly increased the intermolecular spacings, while resin infiltration and resin polymerization each resulted in shrinkage of all the spacings monitored. Critical-point drying for SEM specimens resulted in considerable shrinkage in all three spacings, but was still preferable to air drying, which caused reduction in the order of the fibril packing, resulting in loss of the interfibrillar X-ray pattern. Perhaps the most drastic effect was caused by post-fixation in osmium tetroxide, which resulted in loss of the intermolecular pattern, and also increased the amount of shrinkage in the interfibrillar spacings and the D-periodicity which occurred during later stages of processing.     

Histochemical and ultrahistochemical localization of heavy metals in calf organs

Cadmium, zinc, selenium, and copper were administered, singly or in combination, orally or subcutaneously. Experiment I included 32 calves of both sexes; six received Cd (two groups), Zn, Cd, and Zn, and Cd and Se (two groups) and one group was a control. In Experiment II (21 bulls), three were given Cd, Cd, and Cu, and Cd and Zn, respectively, and one group was a control. For light microscopy, in Experiment I the highest amounts of silver granules were present in the samples of liver, small intestine, and vesicular gland of all the exposed groups; in Experiment II the most affected organs were liver, kidney, and small intestine. For electron microscopy, in Experiment I, after administration of Cd and Zn, the highest amounts of granules were seen in the cytoplasm of hepatocytes and cells of the proximal and distal renal tubules and the lowest amounts were found in glandular cells of the pancreas. Administration of Cd and Se resulted in the presence of large numbers of granules in the nuclei and nucleoli of spermatogonies. In Experiment II, ingestion of Cd and Zn in feed led to the appearance of highest amounts of granules in the nucleoli, nuclei, and cytoplasm of cells in testes, kidneys, and pancreas. Following Cd intake, the highest accumulation of granules was observed in the nucleoli of hepatocytes and cells of the proximal and distal renal tubules. Combined Cd and Cu produced the highest number of granules in cells of the proximal and distal renal tubules and in the nucleoli and nuclei of germinal epithelium.     

Application of 3 × 3 transformation matrix in the correction of three-dimensional AFM image tilts through coordinate transformation

In atomic force microscopy (AFM), it is difficult to maintain samples in a perfect horizontal position on the piezoelectric stage of an AFM to obtain accurate measurement of surface features and to enhance the visual effect of topographic images. Correction of the tilted image therefore requires the use of an appropriate 3 × 3 transformation matrix which will transform the coordinates to simultaneously correct the tilt in both the x-and y-axes due to the three-dimensional nature of AFM images. In this study, application of transformation matrix in three-dimensions for the simultaneous correction of image tilts in both the x- and y-axes by the method of coordinate transformation in three-dimensions is presented. Application of the matrix in simultaneously correcting tilts in the x- and y-axes of an AFM image is implemented by simulating an image assuming there are tilts in both the x- and y-axes. Results of features measurement after transforming the coordinates with the matrix show the efficacy of the matrix. This will conveniently replace the line-by-line analysis done to remove tilts in AFM, which has been the practice long before now. It also has the advantage of correcting images by simultaneously removing tilts in both the x- and y-axes as a veritable replacement of having to separately correcting tilts in images. This matrix can be used to transform coordinates in three-dimensions to obtain accurate step height and linewidth in AFM images.     

Use of primary cell cultures and intact isolated glandular epithelia for X-ray microanalysis

Changes in the elemental composition of cells during isolation of glandular epithelia were studied by electron probe X-ray microanalysis. Fine chopping of rat submandibular gland followed by enzymatic treatment for 15 min caused marked increases in Na and Cl and a decrease in K concentrations in acinar cells. After enzymatic treatment for 50 min, Na, Cl and K concentrations returned to close to the control level. Mechanical disaggregation of the acinar clumps following enzymatic treatment resulted again in minor increases in Na and Cl and a marked decrease in K concentration. Exposure of isolated acini to cholinergic stimulation in vitro resulted in secretion of Cl and K from the acinar cells. Dissection of the sweat gland from human skin caused a decrease in the K/Na ratio. Incubation of the gland for 30–45 min with collagenase gave rise to a gradual decrease in the K/Na ratio. After mechanical separation of the gland into the secretory coil and reabsorptive duct, a further reduction of the K/Na ratio was seen. However, the duct cells had a much lower K/Na ratio and higher Ca concentration than the coil cells. In primary cultures, the K/Na ratios of the coil and duct cells returned to the in situ level. The elemental composition of sweat gland cells incubated in collagenase-containing medium was no different from that in cells incubated in collagenase-free medium. In the intact collagenase-isolated tissue, Cl^? secretion in the coil was elicited by carbachol but not by cAMP, whereas in the duct cells the reverse was the case. In primary cell cultures, Cl^? efflux in both coil and duct cells could be elicited by both carbachol and cAMP. In conclusion, although changes in elemental composition of gland cells during the isolation procedure occur, physiological responses can be detected. When primary cell cultures are used, it should be borne in mind that cultured cells may have physiological properties different from those of the intact tissue.     

Expression of tyrosine hydroxylase and vasopressin in magnocellular neurons of salt-loaded aged rats.

Tyrosine hydroxylase (TH) is expressed in catecholaminergic neurons. However, under certain conditions it is also ectopically expressed in magnocellular neurons of the hypothalamus. To test the hypothesis that this expression of TH is related to the cellular activation of these neurons and/or to the vasopressin (VP) expression, we studied the expression of both TH and VP in control and salt-loaded aged rats. Our results demonstrate that aged rats show a marked TH expression in VP cells which is further increased by osmotic stimulation in the absence of increase in VP synthesis in the supraoptic nucleus. The presence of TH-immunopositive dendritic swellings in the ventral part of this nucleus reveals the high state of plasticity of these neurons. Furthermore, the lack of several actors of catecholamine biosynthesis in these neurons suggests a different role for TH. This study further demonstrates an ectopic expression of TH in hypothalamic neurons of aged rats and a TH expression linked to the activation of VP neurons but unrelated to VP synthesis.     

Immunohistochemistry of neuronal nitric oxide synthase and protein nitration in the striatum of the aged rat

To ascertain the possible implications of the nitric oxide (NO*) producing system in striatal senescence, and by using immunohistochemistry and image-processing approaches, we describe the presence of the enzyme nitric oxide synthase (NOS), the NADPH-diaphorase (NADPH-d) histochemical marker, and nitrotyrosine-derived complexes (N-Tyr) in the striatum of adult and aged rats. The results showed neuronal NOS immunoreactive (nNOS-IR) aspiny medium-sized neurons and nervous fibres in both age groups, with no variation in the percentage of immunoreactive area but a significant decrease in the intensity and in the number of somata with age, which were not related to the observed increase with age of the striatal bundles of the white matter. In addition, NADPH-d activity was detected in neurons with morphology similar to that of the nNOS-IR cells; a decrease in the percentage of area per field and in the number of cells, but an increase in the intensity of staining for the NADPH-d histochemical marker, were detected with age. The number of neuronal NADPH-d somata was higher than for the nNOS-IR ones in both age groups. Moreover, N-Tyr-IR complexes were observed in cells (neurons and glia) and fibres, with a significant increase in the percentage of the area of immunoreaction, related to the increase of white matter, but a decrease in intensity for the aged group. On the other hand, we did not detect the inducible isoform (iNOS) either in adult or in aged rats. Taken together, these results support the contention that NADPH-d staining is not such an unambiguous marker for nNOS, and that increased protein nitration may participate in striatal aging.     

Distribution of sex steroid hormone receptors in the avian brain: functional implications for neural sex differences and sexual behaviors

Developmental and seasonal changes in the production of androgens, estrogens, and progestins seem to control sex-specific differentiation and seasonal changes in appetitive and consummatory sexual behaviors of birds. This results in profound sex differences in the quality (sex-specific) or quantity (sex-typical) of behaviors such as courtship, territoriality, or copulation. Steroids affect the brain by binding to intracellularly located receptors. The same brain areas express androgen, estrogen, and progesterone receptors in male and female brains. Sex differences in these genetically determined patterns occur in the size of neuron populations that intrinsically express sex steroid receptors. Further permanent sex differences are subsequent to degenerative fates of receptor expressing neuron populations during ontogeny. Transient sex differences in receptor expression appear to be due to area-specific up- and down-regulation of receptor levels, reflecting transient changes in the level of circulating steroids, changes in environmental conditions, or in the physiological status of the individuals. In particular, intrinsic sex differences in the expression pattern of sex steroid receptors and steroid-independent regulation of the expression level of these receptors in the brain are limiting mechanisms for gonad-dependent sexual development and activities.     

Mechanotransduction-The relationship between gravity,cells and tensile loading in extracellular matrix

Gravity plays a central role in vertebrate development and evolution. Mechanotransduction involves the tensile tethering of veins and arteries, connections between the epidermis and dermis in skin, tensile stress concentrations that occur at tissue interfaces, cell-cell interactions, cell-collagen fiber stress transfer in extracellular matrix and fluid shear flow. While attention in the past has been directed at understanding the myriad of biochemical players associated with mechanotransduction pathways, less attention has been focused on determining the tensile mechanical behavior of tissues in vivo. Fibroblasts sit on the surface of collagen fibers in living skin and exert a retractile force on the fibers. This retractile force pulls against the tension in collagen fibers in skin. After fibroblast-collagen fiber interactions are altered either by changes in fibroblast adhesion or after formation of cancer associated fibroblasts, and changes in cell junctions, alterations in the retractive force leads to changes in mechanotransduction. The purpose of this paper is to present a model of tensile forces that occur at the fibroblast-collagen fiber interface and how these forces are important in extracellular matrix physiology in health and disease.     

Intestinal and liver changes after chronic ketamine and ketamine plus alcohol treatment

The effects of long‐term chronic ketamine treatment on the intestine and the liver were studied in the ICR mice which had daily intraperitoneal injection of ketamine at 30mg/kg per day for 7 months. The intestine showed no significant pathology after treatment but had a decrease of the positive sites of proliferative cell nuclear antigen in the mucosae of the intestines after ketamine and ketamine plus alcohol (added in the last month) treatment. No significant apoptosis (via TUNEL) nor necrosis (via lactic acid dehydrogenase) was detected in the intestines of all control and ketamine‐treated groups, with the exception of an increase of lactic acid dehydrogenase in the mucosae of the intestines of the ketamine plus alcohol group. In the liver, loss of glycogen was observed in animals after ketamine and ketamine plus alcohol treatment, in addition to the pathology reported in a previous work. The decrease in quantity of glycogen in the liver reflected either a failure of glycogen synthesis from glucose or an increase of glycogenolysis in the liver. Microsc. Res. Tech. 75:1170–1175, 2012. © 2012 Wiley Periodicals, Inc.     

The Ralph knife in practice

Long edge glass knives, ‘Ralph knives’, were produced in an LKB Histo KnifeMaker. The edge angles were measured by light microscopy; depending on the breaking conditions the angles varied between 12° and 58°, as measured close to the edge in the mid-portion of the knives. Hackle marks were more common in the left portion of the edge than in the middle and right portions. Some of the knives were used for cutting sections from urinary bladder tumours embedded in paraffin or in glycol methacrylate. Following microtomy the sections were allowed to stretch on a water surface; this procedure resulted in an increased width of the plastic sections, whereas the paraffin sections were not affected. The compression of the whole sections averaged 15% in the paraffin sections and 11% in the plastic sections; in both cases the compression factors were positively correlated to the angle of the knife edge. Smaller compression factors were found for the cell nuclei in the embedded tissue.     

Comparison of transsynaptic viral labeling of central nervous system structures from the uterine horn in virgin, pregnant, and lactating rats

Using the transneuronal viral tracing method, the central nervous system (CNS) connections of the uterine horn were studied in virgin, pregnant, and in lactating rats. The frequency of viral labeling in the brain and the distribution of virus-infected neurons from the uterine horn were compared among groups. There was a marked difference in the frequency of viral labeling in the brain stem. In virgin rats more than half of the brain stems (5 out of 9) were labeled. In contrast, in pregnant animals viral-labeled neurons were detected in only a few cases (3 out of 16) and almost each brain stem of the lactating group was labeled (12 out of 13). A similar, less marked difference was observed in the hypothalamus. The pattern of distribution of infected neurons was similar in each group. In the brain stem, the nucleus of the solitary tract, dorsal motor nucleus of the vagus, area postrema, gigantocellular and paragigantocellular nucleus, ventrolateral medulla, A5 cell group, and caudal raphe nuclei were the most frequently labeled structures. In the diencephalon, viral-infected neurons were detected primarily in the hypothalamic paraventricular nucleus. The telencephalon was devoid of infected cells. Data suggest that the CNS control of the uterine horn varies depending on reproductive status. The low frequency of brain labeling in pregnant rats may be related to the almost complete lack of sympathetic fibers in the uterus prior to parturition and the very high frequency of labeling in lactating animals to the postpartum hyperinnervation of the uterus.     

五自由度机器人的结构设计及其控制

介绍了串联五自由度机器人的机械结构和机器人控制系统的设计.文中设计的五自由度机器人既可以用于实际生产,也可以用于教学和科研.用于实际生产既可以执行点位控制下的操作,也可以执行连续轨迹控制下的操作;用于教学和科研时,该机器人可用于机器人结构分析、机构运动学分析及机器人控制系统的教学演示和实验操作.它具有结构简单、操控方便等特点,而且可以做进一步的研发.     

Role of nerve growth factor in the olfactory system

Olfactory neurons are unique in the mammalian nervous system because of their capacity to regenerate in adult animals. It has been shown that olfactory receptor cells located in the olfactory epithelium are replaced on a continuous basis and in response to injury throughout the life span of most species. NGF, which is one of the neurotrophic factors, is present in many areas of the central and peripheral nervous system. It has been shown that NGF in the olfactory bulb plays a role in the survival of cholinergic neurons in the horizontal limb of the diagonal band (HDB). Recent studies of NGF in the olfactory bulb suggest that it is involved in the development, maintenance, and regeneration of olfactory receptor cells. In this study, we review reports examining the relationship between NGF in the olfactory bulb and neuronal regeneration and development in the mammalian olfactory systems. Low- and high-affinity NGF receptor immunoreactivity is markedly expressed during regeneration and at different stages of development in the mouse olfactory system. This level of immunoreactivity is no longer present after completion of regeneration and at maturation. Other findings indicate that NGF injected into the olfactory bulb is transported retrogradely to the olfactory epithelium. It has also been shown that continuous anti-NGF antibody injection into the olfactory bulb causes degeneration and olfactory dysfunction. Administration of NGF directory into nasal cavity results in an increase in the expression of olfactory marker protein within the olfactory epithelium in axotomized rats. These findings suggested that the presence of NGF in the olfactory bulb plays an essential role in regeneration, maintenance, and development in the olfactory system of mammals.     

Sexual differences and effect of photoperiod on melatonin receptor in avian brain

Several data suggest that melatonin may influence avian reproduction by acting at the level of the hypothalamic-hypophisial-gonadal axis, and/or on neural circuits controlling reproductive behaviours. The action of melatonin is exerted through specific receptors whose distribution and pharmacological properties have been extensively investigated. This review will focus on the distribution, sexual dimorphism, and dependence upon the photoperiod of melatonin binding sites in avian species with a special emphasis on Japanese quail. Melatonin receptors are widely distributed in avian brain. They are mostly present in the visual pathways of all the investigated species and in the song controlling nuclei of oscine birds. Sexual dimorphism of melatonin binding sites (higher density in males than in females) was detected in some telencephalic nuclei of songbirds, in the visual pathways, and in the preoptic area of quail. The last region plays a key role in the activation of male quail copulatory behaviour and it hosts a large population of gonadotropin-releasing hormone-containing neurons. Sexual dimorphism of melatonin-binding sites in the above-mentioned regions suggests a differential role for this hormone in the modulation of visual perception, gonadotropin production, and seasonally activated behaviours in male and female quail. Further studies are necessary to understand interrelationships among photic cues, gonadal steroids, density, and sexually dimorphic distribution of melatonin receptors.     

Vanadocytes, cells hold the key to resolving the highly selective accumulation and reduction of vanadium in ascidians

Since Henze discovered vanadium in the blood (or coelomic) cells of an ascidian in 1911, this unusual phenomenon has attracted the interest of many investigators. The highest concentration of vanadium (350 mM) in the blood cells of Ascidia gemmata, which belongs to the suborder Phlebobranchia, is 10(7) times higher than that in seawater. Of the approximately 10 types of blood cells, a combination of cell fractionation and neutron-activation analysis revealed that the signet ring cells were the true vanadocytes. In the vanadocytes, 97.6% of the vanadium is in the +3 oxidation state (III). The extremely low pH of 1.9 found in vanadocytes suggests that protons, concentrated by an H(+)-ATPase, might be linked to the accumulation of vanadium energetically. The antigen recognized by a monoclonal antibody, S4D5, prepared to identify vanadocytes, was determined to be 6-PGDH in the pentose phosphate pathway. NADPH produced in the pentose phosphate pathway in vanadocytes is thought to participate in the reduction of vanadium(V) to vanadium(IV). During embryogenesis, a vanadocyte-specific antigen first appears in the body wall at the same time that significant accumulations of vanadium become apparent. Three different vanadium-associated proteins (VAPs) were extracted from the blood cells of vanadium-rich ascidians. These are 12.5, 15, and 16 kDa in size and are associated with vanadium in an approximate ratio of 1:16. The cDNA encoding the 12.5 and 15 kDa VAPs was isolated and the proteins encoded were found to be novel. Further biochemical and biophysical characterization of the VAPs is in progress.