Polymer Assembly Encapsulation of Lanthanide Nanoparticles as Contrast Agents for In Vivo Micro-CT.

Despite recent technological advancements in microcomputed tomography (micro-CT) and contrast agent development, preclinical contrast agents are still predominantly iodine-based. Higher contrast can be achieved when using elements with higher atomic numbers, such as lanthanides; lanthanides also have X-ray attenuation properties that are ideal for spectral CT. However, the formulation of lanthanide-based contrast agents at the high concentrations required for vascular imaging presents a significant challenge. In this work, we developed an erbium-based contrast agent that meets micro-CT imaging requirements, which include colloidal stability upon redispersion at high concentrations, evasion of rapid renal clearance, and circulation times of tens of minutes in small animals. Through systematic studies with poly(ethylene glycol) (PEG)-poly(propylene glycol), PEG-polycaprolactone, and PEG-poly(l-lactide) (PLA) block copolymers, the amphiphilic block copolymer PEG114-PLA53 was identified to be ideal for encapsulating oleate-coated lanthanide-based nanoparticles for in vivo intravenous administration. We were able to synthesize a contrast agent containing 100 mg/mL of erbium that could be redispersed into colloidally stable particles in saline after lyophilization. Contrast enhancement of over 250 HU was achieved in the blood pool for up to an hour, thereby meeting the requirements of live animal micro-CT.

Preparation and Characterization of Lanthanum-Incorporated Hydroxyapatite Coatings on Titanium Substrates.

Titanium (Ti) has been widely used in clinical applications for its excellent biocompatibility and mechanical properties. However, the bioinertness of the surface of Ti has motivated researchers to improve the physicochemical and biological properties of the implants through various surface modifications, such as coatings. For this purpose, we prepared a novel bioactive material, a lanthanum-incorporated hydroxyapatite (La-HA) coating, using a dip-coating technique with a La-HA sol along with post-heat treatment. The XRD, FTIR and EDX results presented in this paper confirmed that lanthanum was successfully incorporated into the structure of HA. The La-HA coating was composed of rod-like particles which densely compacted together without microcracks. The results of the interfacial shear strength test indicated that the incorporation of lanthanum increased the bonding strength of the HA coating. The mass loss ratios under acidic conditions (pH=5.5) suggested that the La-HA coatings have better acid resistance. The cytocompatibility of the La-HA coating was also revealed by the relative activity of alkaline phosphatase, cellular morphology and cell proliferation assay in vitro. The present study suggested that La-HA coated on Ti has promising potential for applications in the development of a new type of bioactive coating for metal implants.

Differences in gastrointestinal calcium absorption after the ingestion of calcium-free phosphate binders.

Both calcium-containing and noncalcium-containing phosphate binders can increase gastrointestinal calcium absorption. Previously, we observed that lanthanum carbonate administration to rats with renal failure is not associated with increased calciuria. Additionally, lanthanum carbonate treatment in dialysis patients has been associated with a less pronounced initial decrease in serum parathyroid hormone compared with other phosphate binders. For 8 days, male Wistar rats received a diet supplemented with 2% lanthanum carbonate, 2% sevelamer, 2% calcium carbonate, or 2% cellulose. Calciuria was found to be increased in animals with normal renal function treated with sevelamer or calcium carbonate but not with lanthanum carbonate. In animals with renal failure, cumulative calcium excretion showed similar results. In rats with normal renal function, serum ionized calcium levels were increased after 2 days of treatment with sevelamer, while calcium carbonate showed a smaller increase. Lanthanum carbonate did not induce differences. In animals with renal failure, no differences were found between sevelamer-treated, calcium carbonate-treated, and control groups. Lanthanum carbonate, however, induced lower ionized calcium levels within 2 days of treatment. These results were confirmed in normal human volunteers, who showed lower net calcium absorption after a single dose of lanthanum carbonate compared with sevelamer carbonate. In conclusion, these two noncalcium-containing phosphate-binding agents showed a differential effect on gastrointestinal calcium absorption. These findings may help to improve the management of calcium balance in patients with renal failure, including concomitant use of vitamin D.

Antibacterial fluorescent nano-sized lanthanum-doped carbon quantum dot embedded polyvinyl alcohol for accelerated wound healing.

Bacteria is one of the main culprits that cause human diseases and pose long-term challenges to people's health. Rare earth elements have unique antibacterial advantages, but little research is available. In this paper, we reported an antibacterial composite film based on lanthanum-doped carbon quantum dot nanoparticles (La@N-P-CQDs) and polyvinyl alcohol (PVA) film for fluorescence of antibiotics and accelerating wound healing. PVA/La@N-P-CQDs composite film presented excellent hydrophilicity, biocompatibility, fluorescence intensity, and antibacterial effects. The antibacterial activity of La@N-P-CQDs was evaluated by employing antibacterial assay using Escherichia coli (E.coli)and Staphylococcus aureus (S.aureus) in vitro. La@N-P-CQDs showed enhanced antibacterial activity compared with N-P-CQDs. Moreover, the PVA/La@N-P-CQDs composite film with 0.5 mg/mL La@N-P-CQDs showed better antibacterial capability and wound healing performance than PVA and PVA/N-P-CQDs films in bacterial adhesion experiment. PVA/La@N-P-CQDs composite film could be used for wound dressing in vivo experiment and had no side effects on major organs in mice. The antibacterial composite film significantly promoted in vivo wound healing process because of its multifunctional properties. Therefore, it was an excellent candidate for wound dressing.

Improving the valence self-reversible conversion of cerium nanoparticles on titanium implants by lanthanum doping to enhance ROS elimination and osteogenesis.

Equipped with anti-oxidative properties, cerium oxide nanoparticles (CNPs) are gradually being adopted over the years in the field of oxidative stress research. However, the effects of CNPs may be diminished when under the influence of prolonged and substantially elevated levels of oxidative stress. Therefore, it is imperative to enhance the efficacy of CNPs to resist oxidative stress. In this study, our approach involves the fabrication of titanium surface CNPs coatings doped with different concentrations of lanthanum ions (La3+) and the investigation of their local anti-oxidative stress potential. The physicochemical characterization showed that the La-CNPs groups had a substantial increase in the generation of oxygen vacancies within the CNPs structure with the increase of La doping concentration. In vitro findings proofed that the cytocompatibility of different La-CNPs coatings showed a trend of increasing first and then decreasing with the increase of La doping concentration under oxidative stress microenvironment. Among these groups, the 30 % La-CNPs group presented the best cell proliferation and osteogenic differentiation which could activate the FoxO1 pathway, then upregulated the expression of SOD1 and CAT, and finally resulted in the inhibition of ROS production. In vivo results further confirmed that the 30 % La-CNPs group showed significant osteogenic effects in two rat models (osteoporosis and diabetes models). In conclusion, we believe that the 30 % La-CNPs coating holds promising potential for its implant applications in patients with oxidative stress-related diseases.

La3+/Sr2+ Dual-Substituted Hydroxyapatite Nanoparticles as Bone Substitutes: Synthesis, Characterization, In Vitro Bioactivity and Cytocompatibility.

The present study aims to synthesize biocompatible and bioactive lanthanum (La3+)/strontium (Sr2+) dual ion doped hydroxyapatite (HA) nanomaterials by sol-gel method. The discrete substitution of La3+ and Sr2+ in pure HA enhances the osteoconductivity. The co-substitution of various La3+ concentrations (0.03, 0.06 and 0.1 M) regulates the physical and In Vitro properties. The study also investigates the effect of La3+/Sr2+ substituents on the crystalline property, microstructure, photoluminescence and In Vitro bioactivity of HA samples. La3+/Sr2+ co-substitution decreases the crystallite size of HA without any significant distortion of the crystal structure. In addition, the dual ions doping influences nanoparticles morphology by reducing the particle size from 75 to 20 nm. The In Vitro bioactivity tests for the La3+/Sr2+ co-substituted HA confirm the osteoconductive boneapatite generating capacity. Bactericidal tests against Staphylococcus aureus and Pseudomonas aeruginosa stains show better resistivity of La3+/Sr2+-HA samples. To authenticate the biocompatibility and antimicrobial activity of the synthesized La3+/Sr2+ dual doped HA nanoparticles for bone implant applications, different tests like cell viability and toxicity were conducted using human lung A549 cells.

Magnetic lanthanum-doped hydroxyapatite/chitosan scaffolds with endogenous stem cell-recruiting and immunomodulatory properties for bone regeneration.

Generally, the addition of exogenous stem cells and host-to-scaffold immune responses restricts the clinical applications of hydroxyapatite (HA)/chitosan (CS) scaffolds for bone regeneration. To achieve "facilitated endogenous tissue engineering", magnetic M-type hexagonal ferrite (SrFe12O19) nanoparticles were incorporated into bone scaffolds to recruit endogenous stem cells. Then, lanthanum incorporation was utilized to regulate host-to-scaffold immune responses and to provide a pro-regenerative environment for recruited endogenous stem cells. Here, we first fabricated and characterized magnetic lanthanum-doped HA/CS scaffolds. The MLaHA/CS scaffolds were demonstrated to be effective at recruiting rat bone marrow mesenchymal stem cells (rBMSCs) and modulating host-to-scaffold immune responses by promoting macrophage polarization into the anti-inflammatory phenotype (M2) in vitro. By further examining the underlying mechanism, we found that MLaHA/CS scaffolds could promote the osteogenic differentiation of rBMSCs by upregulating the phosphorylation of the Smad 1/5/9 pathway. When MLaHA/CS scaffolds were implanted into rat calvarial defects, the incorporation of magnetic nanoparticles and lanthanum significantly promoted the new bone regeneration, as revealed by micro-CT assays and histological staining. Our findings suggest that MLaHA/CS shows great potential for use as a cell-free and biocompatible scaffold for bone regeneration.

Calcium alginate template-mineral substituted hydroxyapatite hydrogel coated titanium implant for tibia bone regeneration.

Osteogenic differentiation is great significance for improving the bone regeneration. Present study evaluates the osteogenic ability of lanthanum (La3+) and silicate (SiO44-) substituted hydroxyapatite (MHAP) - polymeric composite coated surface treated titanium (Ti) implant. The bio-ceramic MHAP was synthesized by hydrothermal process with assistance of calcium alginate template. For enhance the hydrophilicity, the polymer poly (vinyl pyrrolidone) (PVP) was included in the composite by ultra-sonication method. The negative zeta potential value -9.97 mV of Ca-alg/ La, Si-HAP was observed after the incorporation of PVP in the matrix. Incorporation of minerals and PVP polymer was confirmed and analyzed by Energy Dispersive X-ray analysis (EDX), Fourier Transform Infra-Red spectroscopy (FT-IR) and Electron Microscopy techniques. A compact coating of the composite with the thickness of 448 nm on Ti surface was achieved by Electrophoretic deposition (EPD) method. The in-vitro MTT assay method and alkaline phosphate ALP activity (94% and 0.94 a.u respectively for the optimized composite) were utilized to determine the cell viability and differentiation on human Bone Marrow-Derived Stem Cells (hBMSCs). The osteogenic ability of bio-composite coated Ti in hBMSCs and in-vivo rat model has strongly suggests the fabricated Ti plate with bio-composite coatings can act as promising biomaterial for orthopedics.

Role of phi cells and the endodermis under salt stress in Brassica oleracea.

Phi cell layers were discovered in the 19th century in a small number of species, including members of the Brassicaceae family. A mechanical role was first suggested for this structure; however, this has never been demonstrated. The main objective of the present work was to analyse the ultrastructure of phi cells, their influence on ion movement from the cortex to the stele, and their contribution to salt stress tolerance in Brassica oleracea. Transmission electron microscopy and X-ray microanalysis studies were used to analyse the subcellular structure and distribution of ions in phi cells and the endodermis under salt stress. Ion movement was analysed using lanthanum as an apoplastic tracer. The ultrastructural results confirm that phi cells are specialized cells showing cell wall ingrowths in the inner tangential cell walls. X-ray microanalysis confirmed a build-up of sodium. Phi thickenings were lignified and lanthanum moved periplasmically at this level. To the best of our knowledge, this is the first study reporting the possible role of the phi cells as a barrier controlling the movement of ions from the cortex to the stele. Therefore, the phi cell layer and endodermis seem to be regulating ion transport in Brassica oleracea under salt stress.

Blood compatibility improvement of titanium oxide film modified by doping La(2)O(3).

La(2)O(3) doped titanium oxide (TiO(2)) films with different concentration were deposited by means of the Radio-Frequency magnetron sputtering technique. The microstructure and surface properties of TiO(2) films were characterized by X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS) and contact angle test. The blood compatibility of the specimens was evaluated by tests of platelet adhesion. Results show that pure rutile phase is formed in doped samples and La(2)O(3) incorporation significantly improves the wettability and hemocompatibility of TiO(2) films. Our studies demonstrate that La(2)O(3) doped TiO(2) films are potentially useful biomaterials with good blood compatibility.

Lanthanum-silicate-substituted apatite synthesized by fast mechanochemical method: Characterization of powders and biocoatings produced by micro-arc oxidation.

Lanthanum-silicate substituted apatite with equal concentrations of the substituents in the range of 0.2-6.0 mol were produced by a fast method - mechanochemical synthesis. This method makes it possible to synthesize a nanosized single-phase product by activating reaction mixtures containing CaHPO4, CaO, La(OH)3 and SiO2·H2O for 25-30 min in AGO-2 and AGO-3 planetary mills. The structure of the apatites was investigated by the FTIR and XRD methods. It was found that the synthesized samples with substituent concentrations up to 2 mol are substituted oxy-hydroxyapatites, at higher concentrations, they are substituted oxyapatites. The mechanochemically synthesized apatite with a substituent concentration of 0.5 mol was used for depositing biocoatings on titanium substrates by the micro-arc oxidation method. The structure of the coatings is mainly amorphous. In vitro biological tests demonstrated high biocompatibility of the coatings and the absence of cytotoxic action on mesenchymal stem cells.

The controllable lanthanum ion release from Ca-P coating fabricated by laser cladding and its effect on osteoclast precursors.

Several studies have suggested that rare earth oxides can improve properties of bioceramic coating, and bone resorption of osteoclast can be inhibited by rare earth ion releasing certain concentration. However, the effects of lanthanum ion (La3+) released from Ca-P coating on osteoclast precursors is not clear. In this work, La2O3-doped gradient bioceramic coatings were fabricated on Ti alloy (Ti-6Al-4V) by laser cladding with mixed powders of CaHPO4·2H2O, CaCO3 and La2O3. And the bioactivity, mechanical properties and the La3+ release from coating were investigated in vitro. Human osteosarcoma cells (MG63) were used as a cell model to evaluate the biocompatibility of coatings. Mouse macrophage RAW264.7 cells were cultured on coatings to study the effect of La3+ release from Ca-P coating on osteoclast precursors. The XRD results reveal that the amount of HA + TCP reaches maximum (2θ = 32-33°) when the content of La2O3 is 0.6 wt%, and the proliferation of MG63 cells is up to highest value, which indicates that compared with other groups, the bioceramic coating with 0.6 wt% La2O3 is of best biocompatibility. Furthermore, the differentiation of RAW264.7 cells into osteoclast could be inhibited by controllable releasing La3+ from Ca-P coating when soaked in SBF, which demonstrates that controllable La3+ release from Ca-P coating is an effective method to prevent osteoclast formation. And a prospective therapy is provided to cure the disease of wear debris in replacement of artificial joint.

Metal ion-dependent, reversible, protein filament formation by designed beta-roll polypeptides.

BACKGROUND: A right-handed, calcium-dependent beta-roll structure found in secreted proteases and repeat-in-toxin proteins was used as a template for the design of minimal, soluble, monomeric polypeptides that would fold in the presence of Ca2+. Two polypeptides were synthesised to contain two and four metal-binding sites, respectively, and exploit stacked tryptophan pairs to stabilise the fold and report on the conformational state of the polypeptide. RESULTS: Initial analysis of the two polypeptides in the presence of calcium suggested the polypeptides were disordered. The addition of lanthanum to these peptides caused aggregation. Upon further study by right angle light scattering and electron microscopy, the aggregates were identified as ordered protein filaments that required lanthanum to polymerize. These filaments could be disassembled by the addition of a chelating agent. A simple head-to-tail model is proposed for filament formation that explains the metal ion-dependency. The model is supported by the capping of one of the polypeptides with biotin, which disrupts filament formation and provides the ability to control the average length of the filaments. CONCLUSION: Metal ion-dependent, reversible protein filament formation is demonstrated for two designed polypeptides. The polypeptides form filaments that are approximately 3 nm in diameter and several hundred nm in length. They are not amyloid-like in nature as demonstrated by their behaviour in the presence of congo red and thioflavin T. A capping strategy allows for the control of filament length and for potential applications including the "decoration" of a protein filament with various functional moieties.

Effect of lanthanum ions (La3+) on ferritin-regulated antioxidant process under PEG stress.

The physiological effects of lanthanum(III) ions on the ferritin-regulated antioxidant process were studied in wheat (Triticum aestivum L.) seedlings under polyethylene glycol (PEG) stress. Treatment with 0.1 mM La3+ resulted in increased levels of chlorophyll, carotenoid, proline, ascorbate, and reduced glutathione. The activities of superoxide dismutase, catalase, ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductase, glutathione reductase, and peroxidase were also increased after La3+ treatment. Treatment with La3+ seems to enhance the capacity of the reactive oxygen species scavenging system, affect the Fe2+ and Fe3+ electron-transfer process in ferritin, and restrain the formation of hydroxyl radical (OH.), alleviating the oxidative damage induced by PEG stress.

The involvement of parenchymal, Kupffer and endothelial liver cells in the hepatic uptake of intravenously injected liposomes. Effects of lanthanum and gadolinium salts.

125I-labeled albumin or poly(vinyl pyrrolidone) encapsulated in intermediate size multilamellar or unilamellar liposomes with 30-40% of cholesterol were injected intravenously into rats. In other experiments liposomes containing phosphatidyl[Me-14C]choline was injected. 1 h after injection parenchymal or non-parenchymal cells were isolated. Non-parenchymal cells were separated by elutriation centrifugation into a Kupffer cell fraction and an endothelial cell fraction. From the measurements of radioactivities in the various cell fractions it was concluded that the liposomes are almost exclusively taken up by the Kupffer cells. Endothelial cells did not contribute at all and hepatocytes only to a very low extent to total hepatic uptake of the 125I-labels. Of the 14C-label, which orginates from the phosphatidylcholine moiety of the liposomes, much larger proportions were recovered in the hepatocytes. A time-dependence study suggested that besides the involvement of phosphatidylcholine exchange between liposomes and high density lipoprotein, a process of intercellular transfer of lipid label from Kupffer cells to the hepatocytes may be involved in this phenomenon. Lanthanum or gadolinium salts, which effectively block Kupffer cell activity, failed to accomplish an increase in the fraction of liposomal material recovered in the parenchymal cells. This is compatible with the notion that liposomes of the type used in these experiments have no, or at most very limited, access to the liver parenchyma following their intravenous administration to rats.

La(3+) and Gd(3+) induce shape change of giant unilamellar vesicles of phosphatidylcholine.

Lanthanides such as La(3+) and Gd(3+) are well known to have large effects on the function of membrane proteins such as mechanosensitive ionic channels and voltage-gated sodium channels, and also on the structure of phospholipid membranes. In this report, we have investigated effects of La(3+) and Gd(3+) on the shape of giant unilamellar vesicle (GUV) of dioleoylphosphatidylcholine (DOPC-GUV) and GUV of DOPC/cholesterol by the phase-contrast microscopy. The addition of 10-100 microM La(3+) (or Gd(3+)) through a 10-microm diameter micropipette near the DOPC-GUV (or DOPC/cholesterol-GUV) triggered several kinds of shape changes. We have found that a very low concentration (10 microM) of La(3+) (or Gd(3+)) induced a shape change of GUV such as the discocyte via stomatocyte to inside budded shape transformation, the two-spheres connected by a neck to prolate transformation, and the pearl on a string to cylinder (or tube) transformation. To understand the effect of these lanthanides on the shape of the GUV, we have also investigated phase transitions of 30 microM dipalmitoylphosphatidylcholine-multilamellar vesicle (DPPC-MLV) by the ultra-sensitive differential scanning calorimetry (DSC). The chain-melting phase transition temperature and the L(beta') to P(beta') phase transition temperature of DPPC-MLV increased with an increase in La(3+) concentration. This result indicates that the lateral compression pressure of the membrane increases with an increase in La(3+) concentration. Thereby, the interaction of La(3+) (or Gd(3+)) on the external monolayer membrane of the GUV induces a decrease in its area (A(ex)), whereas the area of the internal monolayer membrane (A(in)) keeps constant. Therefore, the shape changes of the GUV induced by these lanthanides can be explained reasonably by the decrease in the area difference between two monolayers (DeltaA=A(ex)-A(in)).

Red cell filterability determined using the cell transit time analyzer (CTTA): effects of ATP depletion and changes in calcium concentration.

Cell transit time analysis (CTTA) is a new filtrometric technique for assessing red blood cell deformability by measuring the conductivity change caused by passage of erythrocytes through a polycarbonate filter. Most reported studies to date using CTTA have focused on the transit time (TT), the duration of passage of an individual red cell through a micropore. Bulk flow rate has not been previously measured via CTTA. The use of new enzyme based cleaning solutions make it possible to reduce clogging in micropore filters. Therefore, valid measures of the number of red cell transits per unit time (counts/s: C/S) can now be obtained. We evaluated both parameters, TT and C/S, as indicators of red cell filterability. Our goal was to evaluate the effect of metabolic changes shown by alternative techniques to affect red cell deformability. The two best established factors are changes in intracellular [ATP] and [Ca2+]. ATP depletion produces a very small increase in TT but a very marked decrease in C/S. In contrast, the addition of low concentrations of calcium produces an increase in TT with minimal decrease in C/S. The effects of calcium appear to be complex. The substantial changes in intracellular calcium induced by the ionophore A23187 result in a curvilinear pattern of increase in transit times and reduction in counts per s. Lanthanum, which inhibits egress of intracellular calcium, causes an increase in TT with a drop in C/S. We conclude that CTTA demonstrates the same changes in red cell deformability measurable by alternative filtrometric techniques; however, CTTA furnishes two separate and independent parameters which may be used to evaluate red cell deformability.

Block of tetrodotoxin-resistant Na+ channel pore by multivalent cations: gating modification and Na+ flow dependence.

Tetrodotoxin-resistant (TTX-R) Na(+) channels are much less susceptible to external TTX but more susceptible to external Cd(2+) block than tetrodotoxin-sensitive (TTX-S) Na(+) channels. Both TTX and Cd(2+) seem to block the channel near the "DEKA" ring, which is probably part of a multi-ion single-file region adjacent to the external pore mouth and is involved in the selectivity filter of the channel. In this study we demonstrate that other multivalent transitional metal ions such as La(3+), Zn(2+), Ni(2+), Co(2+), and Mn(2+) also block the TTX-R channels in dorsal root ganglion neurons. Just like Cd(2+), the blocking effect has little intrinsic voltage dependence, but is profoundly influenced by Na(+) flow. The apparent dissociation constants of the blocking ions are always significantly smaller in inward Na(+) currents than those in outward Na(+) current, signaling exit of the blocker along with the Na(+) flow and a high internal energy barrier for "permeation" of these multivalent blocking ions through the pore. Most interestingly, the activation and especially the inactivation kinetics are slowed by the blocking ions. Moreover, the gating changes induced by the same concentration of a blocking ion are evidently different in different directions of Na(+) current flow, but can always be correlated with the extent of pore block. Further quantitative analyses indicate that the apparent slowing of channel activation is chiefly ascribable to Na(+) flow-dependent unblocking of the bound La(3+) from the open Na(+) channel, whereas channel inactivation cannot happen with any discernible speed in the La(3+)-blocked channel. Thus, the selectivity filter of Na(+) channel is probably contiguous to a single-file multi-ion region at the external pore mouth, a region itself being nonselective in terms of significant binding of different multivalent cations. This region is "open" to the external solution even if the channel is "closed" ("deactivated"), but undergoes imperative conformational changes during the gating (especially the inactivation) process of the channel.

Electro-osmosis and the reabsorption of fluid in renal proximal tubules.

The lateral intercellular spaces (LIS) are believed to be the final common pathway for fluid reabsorption from the renal proximal tubule. We postulate that electrogenic sodium pumps in the lateral membranes produce an electrical potential within the LIS, that the lateral membranes bear a net negative charge, and that fluid moves parallel to these membranes because of Helmholtz-type electro-osmosis, the field-induced movement of fluid adjacent to a charged surface. Our theoretical analysis indicates that the sodium pumps produce a longitudinal electric field of the order of 1 V/cm in the LIS. Our experimental measurements demonstrate that the electrophoretic mobility of rat renal basolateral membrane vesicles is 1 micron/s per V/cm, which is also the electro-osmotic fluid velocity in the LIS produced by a unit electric field. Thus, the fluid velocity in the LIS due to electro-osmosis should be of the order of 1 micron/s, which is sufficient to account for the observed reabsorption of fluid from renal proximal tubules. Several experimentally testable predictions emerge from our model. First, the pressure in the LIS need not increase when fluid is transported. Thus, the LIS of mammalian proximal tubules need not swell during fluid transport, a prediction consistent with the observations of Burg and Grantham (1971, Membranes and Ion Transport, pp. 49-77). Second, the reabsorption of fluid is predicted to cease when the lumen is clamped to a negative voltage. Our analysis predicts that a voltage of -15 mV will cause fluid to be secreted into the Necturus proximal tubule, a prediction consistent with the observations of Spring and Paganelli (1972, J. Gen. Physiol., 60:181).

Extracellular Ca2+ increases cytosolic free Ca2+ in freshly isolated rat odontoblasts.

Recent evidence suggests that extracellular Ca2+ may modulate cell function in mineralized tissue. To determine whether dentinogenic cells, in particular, are sensitive to extracellular Ca2+, fura-2 microfluorometry was used to monitor intracellular calcium levels in odontoblasts freshly isolated from rat incisor. In response to applications of 0.5-4.0 mM extracellular calcium (CaCl2), most odontoblasts (84%; 107/128) showed an increase in intracellular calcium. For the majority of these cells (70%; 75/107), the typical response was biphasic; there was an initial, transient increase in intracellular calcium which reached peak levels within 30-50 s and decayed rapidly, followed by a slower (> 300 s) recovery toward basal levels. In general, the response of these cells to calcium was repeatable and the mean calcium concentration for the half-maximal response was approximately 1.3 mM. This effect could be partially blocked by either 200 microM lanthanum, a nonspecific blocker of Ca2+ channels, or 20 microM dantrolene, a potent inhibitor of Ca2+ release from internal stores. Used in combination, lanthanum, and dantrolene nearly abolished the calcium response completely. In addition, this response was sensitive to the dihydropyridine-sensitive calcium channel blocking agent nicardipine (60 microM), indicating a role for voltage-gated calcium channels during these events. These results show that odontoblasts respond to external calcium through mechanisms involving both influx of external calcium as well as release of calcium from internal stores and suggest a role for extracellular calcium in regulating the function of these cells.