An investigation into the effects of stimulators of fluid production on Locusta Malpighian tubule intracellular elemental composition

The intracellular elemental concentrations of Na, K, P, S, Cl and Mg in the type 1 cells of Malpighian tubules of Locusta migratoria L. have been measured using electron probe X-ray microanalysis. The effects of in vitro stimulation with 1 mM cAMP and corpora cardiaca extract (CC-extract) on the elemental concentrations have been quantified. The distribution of elements, particularly Na, K and Cl is not homogeneous in control cells, and concentration gradients exist within the cytoplasm. Dibutyryl-cAMP (DB-cAMP) caused a decrease in [K]_i without disrupting the gradient which increased from the basal to the apical surface, the apical [Na]_i was increased as was the [Cl]_i. In contrast, in vitro application of CC-extract did not cause changes to the intracellular elemental composition as compared with control cells These data are consistent with the interpretation that exogenous cAMP only partially activated the full stimulatory response of Malpighian tubule cells observed with CC-extract. The changes observed in the density and elemental composition of the `dark bodies' in response to DB-cAMP and CC-extract stimulation suggest that these structures have a role in the ionic economy of Malpighian tubule cells. Accepted: 6 April 1999     

Electron probe X-ray microanalysis of cisplatin-induced cell death in rat pheochromocytoma PC12 cells

Several lines of evidence suggest that cisplatin-induced cell death is not always the result of apoptosis. A distinctive feature between apoptosis and necrosis is the alteration in cell volume regulation and ion homeostasis. Here we analyzed the changes in intracellular element content during cell death induced by exposure to therapeutic concentrations of cisplatin in the PC12 cell line. To quantitate Na, Cl and K content, electron probe X-ray microanalysis (EPXMA) was performed in whole freeze-dried cells. We also traced the alterations in morphological features with fluorescence and transmission electron microscopy. EPXMA demonstrated progressive derangement of the absolute intracellular Na, Cl and K contents. Cisplatin-treated cells showed two microanalytical patterns: 1) cells with alterations in elemental content typical of apoptosis, i.e., an increase in intracellular Na and a decrease in intracellular Cl and K, and 2) cells characterized by an increase in Na content and a decrease in K content, with no changes in Cl content. This intracellular profile for Na, Cl, and K was not typical of necrosis or apoptosis. Morphological analysis revealed two cellular phenotypes: 1) cells characterized by a phenotype typical of apoptosis, and 2) cells characterized by a hybrid phenotype combining variable features of apoptosis and necrosis. Taken together, our findings suggest that therapeutic concentrations of cisplatin may cause a hybrid type of cell death characterized by concurrent apoptosis and necrosis in the same individual PC12 cell.     

X-ray microanalysis of cAMP-induced ion transport in cystic fibrosis fibroblasts.

cAMP-induced ion transport in normal and cystic fibrosis (CF) fibroblasts was investigated by X-ray microanalysis. Stimulation with cAMP causes an increase in cellular Na content and a decrease in cellular Cl and K content. No significant difference in response between CF and normal cells was noted. In this respect, fibroblasts differ from epithelial cells, where cAMP-induced Cl- efflux blocked in CF patients. Isoproterenol produced similar changes in Na and K content as cAMP, but did not effect Cl content.     

Elemental Composition and Water Content of Neuron and Glial Cells in the Central Nervous System of the North American Medicinal Leech (Macrobdella decora)

Elemental (Na, P, S, Cl, K, Ca, Mg) composition and water content of neurons and glial cells of the leech (Macrobdella decora) were determined by x-ray microanalysis of frozen hydrated and dried section techniques. Results are reported as elemental mass fractions (mass/mass) and water content as percent mass. Specific cell compartments and cell types had distinct elemental patterns and water content which suggests that chemical composition of specific cell types is unique and may represent an expression of cell differentiation analogous to morphological specialization. Water content of cells was also cell specific and ranged from 55% (neurons) to 90% (vacuolated zone of glial cells). K and Na were present in concentrations greater than predicted by ion-selective microelectrode measurements, indicating that not all the K and Na were simultaneously accessible to such electrodes.     

Furosemide-sensitive Na and K fluxes in human red cells. Net uphill Na extrusion and equilibrium properties

This paper reports experiments designed to find the concentrations of internal and external Na and K at which inward and outward furosemide-sensitive (FS) Na and K fluxes are equal, so that there is no net FS movement of Na and K. The red cell cation content was modified by using the ionophore nystatin, varying cell Na (Nai) from 0 to 34 mM (K substitution, high-K cells) and cell K (Ki) from 0 to 30 mM (Na substitution, high-Na cells). All incubation media contained NaCl (Nao = 130 or 120 nM), and KCl (Ko = 0-30 mM). In high-K cells, incubated in the absence of Ko, there was net extrusion of Na through the FS pathway. The net FS Na extrusion increased when Nai was increased. Low concentrations of Ko (0-6 mM) slightly stimulated, whereas higher concentrations of Ko inhibited, FS Na efflux. Increasing Ko stimulated the FS Na influx (K0.5 = 4 mM). Under conditions similar to those that occur in vivo (Nai = 10, Ki = 130, Nao = 130, Ko = 4 mM, Cli/Clo = 0.7), net extrusion of Na occurs through the FS pathway (180-250 mumol/liter cell X h). The concentration of Ko at which the FS Na influx and efflux and the FS K influx and efflux become equal increased when Nai increased in high-K cells and when Ki was increased in high-Na cells. The net FS Na and K fluxes both approached zero at similar internal and external Na and K concentrations. In high-K cells, under conditions when net Na and K fluxes were near zero, the ratio of FS Na to FS K unidirectional flux was found to be 2:3. In high-K cells, the empirical expression (Nai/Nao)2(Ki/Ko)3 remained at constant value (apparent equilibrium constant, Kappeq +/- SEM = 22 +/- 2) for each set of internal and external cation concentrations at which there was no net Na flux. These results indicate that in the physiological region of concentrations of internal and external Na, K, and Cl, the stoichiometry of the FS Na and K fluxes is 2 Na:3 K. In high-Na cells under conditions when net FS Na and K fluxes were near zero, the ratio of FS Na to FS K unidirectional fluxes was 3:2 (1).(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of Increased Extracellular K on the Elemental Composition and Water Content of Neuron and Glial Cells in Leech CNS

Elemental (Na, Cl, K) and water contents of leech (Macrobdella decora) neurons and glial cells were determined under steady-state exposure to 4, 10, and 20 mM KCl concentrations (bathing media) using x-ray microanalysis for quantitative digital imaging of frozen hydrated and dried cryosections. Effects of furosemide, 5-hydroxytryptamine (5-HT), and ouabain on elemental distribution changes, induced by exposure to 20 mM K, were also determined. Results demonstrated that packet glial cells and neurons accumulated substantial amounts of K that appeared evenly distributed throughout the cytoplasm. Cell water content also increased as a function of increased cytoplasmic K so that the net effect was an unchanged wet-weight K concentration (expressed as millimoles per kilogram wet weight). Dry-weight Na and Cl concentration (expressed as millimoles per kilogram dry weight) increased slightly in glial cells; however, because cell water increased, both Na and Cl (wet-weight) concentrations decreased. Neurons, in contrast, had no significant change in either Na or K on a wet-weight basis, so a relatively constant Na/K ratio was maintained despite a small, but significant, increase in K (dry weight) and cell water. These increases, like those in packet glia, were a function of exposure to different concentrations of extracellular space K. These changes were completely abolished by 10(-4) M ouabain. Neither furosemide nor 5-HT appeared to affect neuronal or glial K wet-weight concentrations. These data show that both glial cells and neurons can act as substantial reservoirs for K while maintaining stable K concentrations (by altering cell water content and elemental composition). This process appears to depend on a functioning Na+, K+-ATPase system.     

Biphasic behavior of changes in elemental composition during staurosporine-induced apoptosis

Although the identification of events that occur during apoptosis is a fundamental goal of apoptotic cell death research, little is know about the precise sequence of changes in total elemental composition during apoptosis. We evaluated total elemental composition (Na, Mg, P, Cl, S, and K) in relation to molecular and morphological features in human U937 cells induced to undergo apoptosis with staurosporine, an intrinsic pathway activator. To evaluate total elemental content we used electron probe X-ray microanalysis to measure simultaneously all elements from single, individual cells. We observed two phases in the changes in elemental composition (mainly Na, Cl and K). The early phase was characterized by a decrease in intracellular K (P < 0.001) and Cl (P < 0.001) content concomitant with cell shrinkage, and preceded the increase in proteolytic activity associated with the activation of caspase-3. The later phase started with caspase-3 activation, and was characterized by a decrease in the K/Na ratio (P < 0.001) as a consequence of a significant decrease in K and increase in Na content. The inversion of intracellular K and Na content was related with the inhibition of Na⁺/K⁺ ATPase. This later phase was also characterized by a significant increase (P < 0.001) in intracellular Cl with respect to the early phase. In addition, we found a decrease in S content and an increase in the P/S ratio. These distinctive changes coincided with chromatin condensation and DNA fragmentation. Together, these findings support the concept that changes in total elemental composition take place in two phases related with molecular and morphological features during staurosporine-induced apoptosis.     

Quantitative X-ray microanalysis of the morula cell of the blood of the ascidian Halocynthia papillosa (Stolidobranchiata)

Summary The elemental composition of the morula cell of Halocynthia papillosa blood was studied by X-ray microanalysis with respect to the possible iron accumulation in this cell type. We found various amounts of Na, Mg, P, S, Cl, K, Ca, Fe and Br in the cytoplasm, nucleus and vacuoles. With the exception of a few cells, Ca, Fe and Br were not detected. Thus, the morula cells of the studied species are not iron-rich cells.     

Model of ion transport regulation in chloride-secreting airway epithelial cells. Integrated description of electrical, chemical, and fluorescence measurements.

An electrokinetic model was developed to calculate the time course of electrical parameters, ion fluxes, and intracellular ion activities for experiments performed in airway epithelial cells. Model variables included cell [Na], [K], [Cl], volume, and membrane potentials. The model contained apical membrane Cl, Na, and K conductances, basolateral membrane K conductance, Na/K/2 Cl and Na/Cl symport, and 3 Na/2 K ATPase, and a paracellular conductance. Transporter permeabilities and ion saturabilities were determined from reported ion flux data and membrane potentials in intact canine trachea. Without additional assumptions, the model predicted accurately the measured short-circuit current (Isc), cellular conductances, voltage-divider ratios, open-circuit potentials, and the time course of cell ion composition in ion substitution experiments. The model was used to examine quantitatively: (a) the effect of transport inhibitors on Isc and membrane potentials, (b) the dual role of apical Cl and basolateral K conductance in cell secretion, (c) whether the basolateral symporter requires K, and (d) the regulation of apical Cl conductance by cAMP and Ca-dependent signaling pathways. Model predictions gave improved understanding of the interrelations among transporting systems and in many cases gave surprising predictions that were not obvious without a detailed model. The model developed here has direct application to secretory or absorptive epithelial cells in the kidney thick ascending limb, cornea, sweat duct, and intestine in normal and pathophysiological states such as cystic fibrosis and cholera.     

X-ray microanalysis of the Malpighian tubules of the black field cricket Teleogryllus oceanicus: the roles of Na K ATPase and the Na K 2Cl cotransporter

Both main and distal segments of the Malpighian tubules were sensitive to ouabain and furosemide but in different ways. Oubain had no effect on secretion rate by the main segment but in the secreted fluid Na(+) concentration increased substantially whereas K(+) decreased. Similarly intracellular elemental Na concentration increased and K decreased. Furosemide decreased the secretion rate of the main segment by 80%. The Na(+) concentration in the secreted fluid increased markedly but K(+) was not affected. Intracellular elemental Na concentration also increased but K was unchanged. In the distal segments both ouabain and furosemide decreased secretion rate by 40% but although ouabain had no effect on the composition of the secreted fluid, furosemide caused a substantial reduction in the concentrations of Mg(2+) and Cl(-) and a substantial increase in Na(+) and K(+) concentrations. The evidence suggests that the main segment contains a Na K ATPase and possibly a Na K 2Cl cotransporter whereas the distal segment may contain a Na K ATPase and a furosemide sensitive Mg(2+) transporter. K(+) entry into the cells of the main segment may be partially effected by a Na K 2Cl cotransporter but may be primarily via Na K ATPase in the distal segment.     

Distribution of Elements in Rat Peripheral Axons and Nerve Cell Bodies Determined by X-Ray Microprobe Analysis

X-ray microprobe analysis was used to determine concentrations (millimoles of element per kilogram dry weight) of Na, P, Cl, K, and Ca in cellular compartments of frozen, unfixed sections of rat sciatic and tibial nerves and dorsal root ganglion (DRG). Five compartments were examined in peripheral nerve (axoplasm, mitochondria, myelin, extraaxonal space, and Schwann cell cytoplasm), and four were analyzed in DRG nerve cell bodies (cytoplasm, mitochondria, nucleus, and nucleolus). Each morphological compartment exhibited characteristic concentrations of elements. The extraaxonal space contained high concentrations of Na, Cl, and Ca, whereas intraaxonal compartments exhibited lower concentrations of these elements but relatively high K contents. Nerve axoplasm and axonal mitochondria had similar elemental profiles, and both compartments displayed proximodistal gradients of decreasing levels of K, Cl, and, to some extent, Na. Myelin had a selectively high P concentration with low levels of other elements. The elemental concentrations of Schwann cell cytoplasm and DRG were similar, but both were different from that of axoplasm, in that K and Cl were markedly lower whereas P was higher. DRG cell nuclei contained substantially higher K levels than cytoplasm. The subcellular distribution of elements was clearly shown by color-coded images generated by computer-directed digital x-ray imaging. The results of this study demonstrate characteristic elemental distributions for each anatomical compartment, which doubtless reflect nerve cell structure and function.     

Chloride channel regulation in secretory epithelia

Patch-clamp techniques were applied for single-channel recording to cultured cells from Cl secretory epithelia: human airway cells and the T84 cell line. Epinephrine or cyclic AMP (cAMP) stimulated single-channel activity in human airway cells during cell-attached recording. Similarly, prostaglandin E2 and cAMP stimulated single-channel activity in T84 cells. Ion substitution experiments with patches in the inside-out configuration indicated greater than 10:1 selectivity for Cl over Na in channels from both cell types, which confirms the identity of these events as Cl channel openings. The Ca ionophore A23187 stimulated these Cl channels to open in both cell types. Human airway cells from patients with cystic fibrosis (CF) did not respond to epinephrine or cAMP, but A23187 treatment elicited Cl channel activity. Changes in bath Ca activity in the inside-out configuration demonstrated that increased Ca could activate cAMP-insensitive Cl channels in CF cells. This indicates that the primary defect in CF is in the regulation of Cl channel opening rather than in conduction of Cl through the channel.     

Effects of Axotomy on Distribution and Concentration of Elements in Rat Sciatic Nerve

X-ray microprobe analysis was used to determine the effects of axotomy on distribution and concentration (millimoles of element per kilogram dry weight) of Na, P, Cl, K, and Ca in frozen, unfixed sections of rat sciatic nerve. Elemental concentrations were measured in axoplasm, mitochondria, and myelin at 8, 16, and 48 h after transection in small-, medium-, and large-diameter fibers. In addition, elemental composition was determined in extraaxonal space (EAS) and Schwann cell cytoplasm. During the initial 16 h following transection, axoplasm of small fibers exhibited a decrease in dry weight concentrations of K and Cl, whereas Na and P increased compared to control values. Similar changes were observed in mitochondria of small axons, except for an early, large increase in Ca content. In contrast, intraaxonal compartments of larger fibers showed increased dry weight levels of K and P, with no changes in Na or Ca concentrations. Both Schwann cell cytoplasm and EAS at 8 and 16 h after injury had significant increases in Na, K, and Cl dry weight concentrations, whereas no changes, other than an increase in Ca, were observed in myelin. Regardless of fiber size, 48 h after transection, axoplasm and mitochondria displayed marked increases in Na, Cl, and Ca concentrations associated with decreased K. Also at 48 h, both Schwann cell cytoplasm and EAS had increased dry weight concentrations of Na, Cl, and K. The results of this study indicate that, in response to nerve transection, elemental content and distribution are altered according to a specific temporal pattern. This sequence of change, which occurs first in small axons, precedes the onset of Wallerian degeneration in transected nerves.     

Ganglioside Treatment Modifies Abnormal Elemental Composition in Peripheral Nerve Myelinated Axons of Experimentally Diabetic Rats

Abstract: Effects of ganglioside administration on elemental composition of peripheral nerve myelinated axons and Schwann cells were determined in streptozotocin-induced diabetic rats and nondiabetic controls. Diabetic rats (50 days after administration of streptozocin) exhibited a loss of axoplasmic K and Cl concentrations in sciatic nerve relative to control, whereas intraaxonal levels of these elements increased in tibial nerve. These regional changes in diabetic rat constitute a reversal of the decreasing proximodistal gradients for K and Cl concentrations that characterize normal peripheral nerve. Treatment of diabetic rats with a ganglioside mixture for 30 days (initiated 20 days after the administration of streptozocin) returned proximal sciatic nerve axoplasmic K and Cl concentrations to control levels, whereas in tibial axons, concentrations of these elements increased further relative to diabetic levels. Also in the ganglioside/diabetic group, mean axoplasmic Na concentrations were reduced and Ca levels were elevated. Mixed ganglioside treatment of nondiabetic rats significantly increased axoplasmic dry weight concentrations of K and Cl in proximal sciatic and tibial axons. Schwann cells did not exhibit consistent alterations in elemental content regardless of treatment group. Changes in elemental composition evoked by ganglioside treatment of diabetic rats might reflect the ability of these substances to stimulate Na⁺,K⁺-ATPase activity and might be related to the mechanism by which gangliosides improve functional deficits in experimental diabetic neuropathy.     

Ionic Permeability and Electrical Potential Differences in Necturus Kidney Cells

The cellular concentrations of Na, K, and Cl have been measured in kidney slices of the amphibian, Necturus maculosus. Permeability coefficients have been determined for Na, K, Cl, Rb, Cs, and choline, from studies both of the uptake of radioactive isotopes and the rate of cell swelling in anisotonic solutions. The results of both methods were found to agree well. Measurements were also made of electrical potential differences across the peritubular face of the kidney cells using bathing solutions in which the electrolyte composition and concentrations could be varied. The data obtained are consistent with a model cell in which the potential difference arises as a result of differences in Na permeability relative to K on the two faces of the cell. The intracellular Na concentration is considered to be regulated by a Na-K coupled pump located at the peritubular face of the cell.     

Electron probe analysis, X-ray mapping, and electron energy-loss spectroscopy of calcium, magnesium, and monovalent ions in log-phase and in dividing Escherichia coli B cells.

The elemental composition of individual cells of rapidly frozen and cryosectioned Escherichia coli B was measured with electron optical microanalytic methods. The Ca content was high (26.2 mmol/kg) in a 10-nm-wide region of the cell envelope. Amounts of cytoplasmic Ca in actively dividing cells were significantly higher (32.6 mmol/kg [dry weight]) than in the log-phase (1.5 mmol/kg) cells. Cellular Mg was 205 mmol/kg (dry weight) and it was uniformly distributed throughout the cell. Cells washed in distilled water before freezing lost monovalent ions (Na, Cl, and K), but the membrane-bound Ca and cellular Mg were not reduced, indicating that cellular Mg and membrane Ca are more tightly bound.     

ELEMENTAL COMPOSITION,CORRELATIONS, AND RATIOS WITHIN A POPULATION OF STAURASTRUM PLANCTONICUM (ZYGNEMATALES): AN X-RAY MICROANALYTICAL STUDY1

Individual cells of Staurastrum planctonicum (Teil.) within a mixed freshwater phytoplankton sample were analyzed by scanning electron microscope X-ray microanalysis to determine their elemental composition. X-ray emission spectra routinely showed clear peaks of P, S, and Cl, plus monovalent (Na, K, and divalent (Mg, Ca) cations. Si and Cu were also present in lower quantities. Concentrations of individual elements (expressed as mmol.kg⁻¹ dry weight) varied widely among cells, with values over the sample population approximating to a normal distribution. Although intracellular anion and cation levels varied considerably, significant correlations occurred between concentrations of monovalent and divalent cations (mean ratio 1.4), major diffusible anions and cations (mean ratio 1,2), and total levels of electropositive and electronegative elements (mean ratio 1.2). The monovalent cations of K and Na occurred at a mean ratio of 1.2 and were not significantly correlated. Concentrations of individual elements (except Si) showed clear positive correlations within the analyses, with 12 highly significant (99% probability) correlations out of 36 possible combinations. Principal factor analysis showed that elemental correlations were determined by two major factors, with two resulting groups of elements—(Na, S, Cl, Ca, Cu) and (Mg, P, K). Statistical relationships between elements followed a clear correlation pattern, which retained its characteristics even when elemental concentrations were expressed per unit P rather than per unit dry weight. Elemental concentrations were closely similar in matching, but not nonmatching, smicells. The statistical pattern of elemental associations noted in Staurastrum parallels that seen in X-ray micro-analytical studies of other algae but differs in detail. This pattern of statistical associations has biological implications in terms of cell compartmentation, characterization of different cell types, and cell interactions with their environment.     

Electrical Potentials and Na, K, and Cl Concentrations in the Vacuole and Cytoplasm of Nitella translucens

The potential differences across the tonoplast and plasmalemmamembranes have been measured in the single cells of Nitellatranslucens, the cells being immersed in an artificial pondwater (composition: NaCl _1.0 mM., KC1 _0.1 mM., CaCl₂, _0.1 mM.).The potential of the cytoplasm is –138 m V with respectto the bathing medium and –18 mV with respect to the vacuole.The concentrations of Na, K, and Cl have been measured in thetwo cell fractions. The concentrations in the flowing cytoplasmare: Na 14 mM., K 119 mM., and Cl 65 mM.; the vacuolar concentrationsare: Na 65 mM., K 75 mM.,and Cl 160 mM. The observed potential differences across the two membranesare compared with the Nernst potentials for all three ions.This analysis shows that all three ions are actively transportedat the plasmalemma: Na is pumped outwards while K and Cl arepumped inwards. At the tonoplast Na is pumped into the vacuolewhile K and Cl are close to electrochemical equilibrium. The inhibitor, ouabain, has no effect on the cell resting potential.     

Kinetic analysis of Na,K-activated adenosine triphosphatase induced by low external K+ in a rat liver cell line

Exposure of ARL 15 cells to medium containing reduced concentrations of K+ (0.65 mM) elicited a 50-100% increase in Na,K-ATPase activity. The inhibition by ouabain of both the basal and the induced enzyme conformed to a single-site model (KI = 1 x 10(-4) M). The low K+-induced increment in Na,K-ATPase activity was accompanied by an equivalent increase in the abundance of Na,K-pump sites estimated by ouabain-stabilized ("back-door") phosphorylation, such that the calculated catalytic turnover number of approximately 8000/min was minimally changed. Comparison of the dependence of ouabain-inhibitable K+ uptake on intracellular Na+ and on extracellular K+ concentrations in control and low K+-treated cells revealed no change in the respective half-maximal stimulatory concentrations for these cations, whereas the maximal rate of active K+ uptake in cells exposed to low external K+ increased by nearly 100%. The derived Hill coefficients for active K+ transport rate were also unchanged by the low K+ treatment (i.e. approximately 1.4 for extracellular K+ and 2.6 for intracellular Na+). Na,K-ATPase activity of basal and low K+-induced cells calculated from the measured maximal Na,K transport rate closely approximated the Na,K-ATPase activity measured enzymatically in unfractionated cell lysates under Vmax conditions, suggesting that all or most of the Na,K-ATPase enzymatic units present in both basal and stimulated states are functionally active. Northern blot analysis of RNA isolated from control cells indicated the presence of the Na,K-ATPase alpha-I isoform of the enzyme which increased by nearly 200% following incubation of the cells in low-K+ medium. By contrast, the alpha-II and alpha-III mRNAs were undetectable in either the basal or low K+-stimulated state. These results indicate that the Na,K-ATPase induced by incubation of ARL 15 cells in low-K+ medium is kinetically and functionally indistinguishable from the basal enzyme, and that only the alpha-I isoform is expressed under control and low-K+ conditions.     

Elemental changes associated with chondrocyte differentiation in rat rib growth plate

Quantitative X-ray microanalysis was under-taken to follow the elemental changes that occur in the process of chondrocyte differentiation. For analysis at the cellular level, semi-thick freeze-dried cryosections of rat rib growth plate cartilage were used. For evaluation of the elemental concentrations at the subcellular level, thin sections of freeze-dried and low temperature vacuum embedded cartilage were analyzed. Levels of Na, P, S, Cl, K, and Ca were determined in the cells and extracellular matrix in different zones of the cartilage--resting, proliferative, and hypertrophic. Proliferative cells had a sodium concentration that was twice that of resting cells, suggesting that Na may play an important role in the regulation of DNA- and protein-synthesis in chondrocytes, A concomitant rise in Na and S concentration occurred between resting zone and proliferative zone cartilage matrix. The high concentrations of Na and K in the matrix are probably due to the high amount of sulfate in proteoglycans which may bind these cations.