Nonsaturable Process Research Articles

Effect of vitamin D3 on duodenal calcium absorption in vivo during early development.

The effect of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] and vitamin D deficiency on duodenal calcium absorption in suckling and weaned rats was determined by an in situ loop technique. In vitamin D-replete (+D) rats, the linear, or nonsaturable, component of calcium absorption was very efficient in 14-day-old pups and decreased with age until 35 days. The saturable component, which was undetectable in 14-day-old pups, became detectable by 18 days of age and increased until 26 days of age. Calcium absorption was not reduced in vitamin D-deficient (-D) 14-day-old pups as compared with +D pups. A high dose of 1,25(OH)2D3 increased the plasma calcium level of +D suckling rats but had no effect on calcium absorption even with milk present in the loop. Weaned -D rats had a reduced saturable component of absorption (P less than 0.01) compared with +D rats. A high dose of 1,25(OH)2D3 significantly increased calcium absorption and plasma calcium levels of -D rats. Our results indicate that during suckling calcium absorption occurs by a process that is insensitive to vitamin D. After weaning both saturable and nonsaturable processes appear to contribute to calcium absorption, and the saturable component can be influenced by vitamin D status or a high dose of 1,25(OH)2D3.

The uptake of ornithine and lysine by isolated hepatocytes and fibroblasts

1. 1. The uptake of ornithine and lysine by isolated hepatocytes and cultured human skin fibroblasts were studied. Both types of cells can accumulate these amino acids via a saturable and a non-saturable process, the latter being active at high substrate concentration. 2. 2. The apparent Km for ornithine and lysine for the saturable process in hepatocytes were 0.26 and 0.94 mM respectively, those values for fibroblasts were found to be 0.43 and 0.57 mM respectively. 3. 3. Fractionation of the cells by the digitonin technique into a cytosolic compartment and a paniculate compartment, showed that a rapid equilibration occurs between the external medium and the cytosolic compartment of hepatocytes and fibroblasts.

Intestinal absorption mechanism of amino-beta-lactam antibiotics. III. Kinetics of carrier-mediated transport across the rat small intestine in situ.

The transport kinetics of amino-beta-lactam antibiotics was studied by an in situ rat small intestinal recirculating perfusion technique. The disappearance rates of the antibiotics from the perfusing luminal solution followed mixed-type kinetics with saturable and nonsaturable processes. The kinetic parameters were determined. Pharmacokinetic analysis of the time courses of luminal disappearance, tissue accumulation, and blood concentration indicated that the transfer of the antibiotics from the in situ luminal solution to tissue is nearly irreversible. On the assumption that the saturable transport process involves a common carrier for these antibiotics, the predicted extents of mutual inhibition using the in situ kinetic parameters were in good agreement with the experimental values for cephalexin and cephradine. The effects of cephalexin and cefadroxil on the absorption of cyclacillin were also consistent with a common transport mechanism. The dipeptides, carnosine and L-phenylalanylglycine markedly inhibited cyclacillin absorption in a competitive fashion. Furthermore, cyclacillin inhibited the absorption of carnosine. The results indicate that the absorption of amino-beta-lactam antibiotics is closely related with that of dipeptides.

Uptake of 17 beta-estradiol and its conjugates by isolated rabbit liver nuclei.

The present study compares the uptake and metabolism of 17 beta-estradiol, 17 beta-estradiol 3-glucoside, and 17 beta-estradiol 3-glucuronide by a highly purified preparation of rabbit liver nuclei. The uptake of the three estrogens was rapid and equilibration was reached within 60 s. The order of uptake was 17 beta-estradiol (64 fmol X mg protein -1) greater than 17 beta-estradiol 3-glucoside (10 fmol X mg protein -1) greater than 17 beta-estradiol 3-glucuronide (6.5 fmol X mg protein -1). Thin-layer chromatography of the estrogens taken up by rabbit liver nuclei indicated the presence of a beta-glucosidase activity associated with the nuclear preparation. The apparent Km value of this enzyme for 17 beta-estradiol 3-glucoside (3.5 microM) was about 10-fold higher when compared with the cytosolic enzyme. The uptake of the three estrogens was linearly proportional to the substrate concentration from 1 to 100 nM. No competition for uptake was observed among the steroids and the presence of diethylstilbestrol did not reduce the uptake of the steroids. These findings suggest that 17 beta-estradiol, 17 beta-estradiol 3-glucoside, and 17 beta-estradiol 3-glucuronide are taken up by nuclei by a nonsaturable diffusion process. The effect of cytosol on the uptake of estrogens by purified nuclei was also investigated. It was observed that cytosol reduced the uptake o 17 beta-estradiol but had little effect on that of its conjugates.

Duodenal and ileal calcium absorption in the rat and effects of vitamin D.

An in situ ligated loop procedure was applied to dissect transmural calcium transport in the intestine into two components, a saturable and a nonsaturable process. The existence of two such processes was confirmed in the duodenum, but ileal calcium transport was devoid of the saturable component. There was a small saturable component in the upper jejunum. The level of CaBP, the vitamin D-dependent cytosolic calcium-binding protein (Mr, approximately or equal to 9,000), corresponded to the magnitude of the saturable component. No CaBP was detected in the ileum. Vitamin D dependence of the saturable component was established by inducing it in the duodenum of vitamin D-deficient animals following intraperitoneal injection of 1,25-dihydroxyvitamin D3. In these same animals, conversely, the ileum did not respond to exogenous 1,25-dihydroxyvitamin D3. This confirms the absence in the ileum of the saturable component of transmural calcium movement and the fact that the nonsaturable component is not vitamin D dependent. Everted sac experiments also showed that duodenal sacs from vitamin D-replete or -repleted animals transported calcium against a chemical gradient, whereas ileal sacs did not. Vitamin D regulation of intestinal calcium absorption thus occurs only in the proximal intestine, even though calcium is absorbed down its chemical gradient all along the small intestine.

The regulation of 3-hydroxy-3-methylglutaryl-CoA reductase activity, cholesterol esterification and the expression of low-density lipoprotein receptors in cultured monocyte-derived macrophages.

Human blood monocytes cultured in medium containing 20% whole serum showed the greatest activity of 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase and [14C]acetate incorporation into non-saponifiable lipids around the 7th day after seeding, the period of greatest growth. Although there was enough low-density lipoprotein (LDL) in the medium to saturate the LDL receptors that were expressed by normal cells at that time, HMG-CoA reductase activity and acetate incorporation were as high in normal cells as in cells from familial-hypercholesterolaemic (FH) patients. Both the addition of extra LDL, which interacted with the cells by non-saturable processes, and receptor-mediated uptake of acetylated LDL significantly reduced reductase activity and increased incorporation of [14C]oleate into cholesteryl esters in normal cells and cells from FH patients ('FH cells'), and reduced the expression of LDL receptors in normal cells. Pre-incubation for 20h in lipoprotein-deficient medium apparently increased the number of LDL receptors expressed by normal cells but reduced the activity of HMG-CoA reductase in both normal and FH cells. During subsequent incubations the same rate of degradation of acetylated LDL and of non-saturable degradation of LDL by FH cells was associated with the same reduction in HMG-CoA reductase activity, although LDL produced a much smaller stimulation of oleate incorporation into cholesteryl esters. In normal cells pre-incubated without lipoproteins, receptor-mediated uptake of LDL could abolish reductase activity and the expression of LDL receptors. The results suggested that in these cells, receptor-mediated uptake of LDL might have a greater effect on reductase activity and LDL receptors than the equivalent uptake of acetylated LDL. It is proposed that endogenous synthesis is an important source of cholesterol for growth of normal cells, and that the site at which cholesterol is deposited in the cells may determine the nature and extent of the metabolic events that follow.

Developmental changes in the mechanisms of duodenal calcium transport in the rat.

Duodenal calcium transport was resolved into a saturable and a nonsaturable process by means of an in situ ligated loop procedure applied to Wistar rats at 3, 12, 19, 24, 30, 40, 60, 110, and 150 days of age. All postweaning animals were males that had been placed on a 1.5% calcium, 1.5% phosphorus semisynthetic diet. Duodenal calcium-binding protein (CaBP) levels were determined at all ages. The newborn rat had no saturable transport component and no CaBP. Its nonsaturable component was very high. With increasing age the saturable component and CaBP varied biphasically, increasing steeply until the animals were about 35 days old; thereafter, each decreased to low but detectable values. The nonsaturable component, on the other hand, decreased in near-linear fashion in the first 35 days; in animals beyond that age it remained invariant. The difference in age dependence between the saturable and nonsaturable components may be considered to constitute additional evidence for the existence of the two transport processes. CaBP and the saturable transport process were highly correlated, further proof that both are vitamin D dependent. Histological studies have revealed the presence of many vacuoles in the intestinal cells of the very young rats; these vacuoles were absent in rats older than 35 days. It is suggested that these vacuoles may be implicated in a pinocytosislike nonsaturable transport that is superimposed on the nonsaturable, non-vitamin D-dependent calcium transport found in all enterocytes.

Horseradish peroxidase transport across adult rabbit jejunum in vitro.

Quantification and functional characteristics of intact protein uptake, metabolic behavior, and transmission across the intestinal wall were examined using horseradish peroxidase (HRP) transport through adult rabbit jejunum. Transepithelial HRP fluxes were determined with a modified Ussing apparatus. In Ringer solution, no significant difference was found between intact HRP fluxes from mucosa to serosa (JHRPm leads to s) and those from serosa to mucosa (JHRPs leads to m) (3.12 +/- 0.58 and 3.48 +/- 0.45 pmol.h-1.cm-2, respectively). In the presence of 10 mM glucose, slight net secretion was noted. The transport mechanism was shown to be sensitive to metabolic inhibitors, colchicine, and ammonia. Intracellular transfer and catabolism were estimated by measuring transepithelial fluxes of tritiated horseradish peroxidase (J[3H]HRP). Net absorption of 3H equivalent HRP (91 +/- 32 pmol.h-1.cm-2) occurred chiefly in the form of tritiated degraded catabolites of 2-4 kilodaltons. Comparison of the transepithelial fluxes of intact and 3H equivalent HRP made it possible to estimate that 97% of the HRP was degraded while crossing the tissue from mucosa to serosa and 88% while crossing from serosa to mucosa. The saturable absorption observed for JHRPm leads to s became a nonsaturable process for J[3H]HRPm leads to s. These results fit the existence of at least two functional pathways for intestinal protein transport. The main route seems to involve endocytosis, with striking intracellular degradation, possibly during passage through the lysosomal system. The HRP that escapes metabolic degradation is transported by an alternative route requiring the structural and metabolic integrity of the epithelial cells. Although this route only accounts for a small fraction of HRP transport, it may be of immunological importance.

Effect of sodium concentration and plasma sugar concentration on hexose absorption by the rat jejunum in vivo. Further evidence of two transport mechanisms.

The sodium-dependency of both the saturable and non-saturable components of glucose and galactose absorption across the rat jejunum in vivo has been determined. The non-saturable process appears to be unaffected by sodium removal but when Na+ concentrations in the lumenal fluid are progressively reduced from 143 mM to 0 mM the Jmax for active absorption is greatly decreased. In a separate study these two components of glucose transport were further investigated using intravenous sugar infusion to modify the transepithelial sugar concentration gradient. When identical glucose concentrations were present in plasma and intestinal lumen, the reduction in glucose absorption was fully accounted for by the elimination of the non-saturable component from the overall absorptive process. Together, these observations can be interpreted as further evidence for the existence of at least two absorption processes in vivo. The implication for analysing results from experimental studies of intestinal sugar absorption in vivo is discussed.

Active transport of nicotine by the isolated choroid plexus in vitro.

In vitro, the transport of [14C]nicotine into the isolated choroid plexus, the anatomical locus of the blood--CSF barrier, was studied. The isolated rabbit choroid plexus accumulated [14C]nicotine by two processes: an active saturable transport process and a nonsaturable process. The [14C]nicotine accumulation process by choroid plexus was not due to binding or intracellular metabolism of the [14C]nicotine. The [14C]nicotine accumulation process in isolated choroid plexus was inhibited by weak bases, including tolazoline and lidocaine, but not by the weak acid probenecid. The accumulation process was decreased 60% by iodoacetate and dinitrophenol and by low temperatures. These results are consistent with previous autoradiographic evidence showing the choroid plexus concentrated [14C]nicotine in vivo, and suggest that the choroid plexus may transfer nicotine between blood and CSF in vivo.

Serotonergic innervation of the lateral hypothalamus: Evidence from synaptosomal uptake studies

Uptake of tritiated serotonin by synaptosomes prepared from rat lateral hypothalamus was examined. Uptake of serotonin into lateral hypothalamic synaptosomes occurred by both saturable and non-saturable processes. The saturable process was a high affinity transport with kinetic parameters that agree closely with those previously reported for serotonin uptake into synaptosomes prepared from other brain regions. Fluoxetine, a selective inhibitor of uptake into serotonergic neurons, was a potent inhibitor of serotonin uptake into lateral hypothalamic synaptosomes. Desipramine and benztropine, noradrenergic and dopaminergic uptake inhibitors respectively, were much less effective. Damage to the ascending serotonergic system, by either electrolytic lesion of the dorsal or median raphe nucleus, or by 5,7 dihydroxytryptamine injections into the midbrain serotonergic pathways, significantly reduced the uptake of serotonin by lateral hypothalamic synaptosomes. Taken together, these data provide further evidence for a serotonergic terminal field within the lateral hypothalamus.

Characterisation of medium-inaccessible adenine nucleotides of rabbit skeletal muscle sarcoplasmic reticulum

ATP and ADP associate irreversibly with isolated sarcoplasmic reticulum vesicles in the presence of cytoplasmic concentrations of ATP, Mg 2+, and Ca 2+ (Aderem, A. A., McIntosh, D. B., and Berman, M. C., 1979, Proc. Nat. Acad. Sci. USA 76, 3622–3626). The nature of these medium-inaccessible adenine nucleotides has been studied from the kinetics of appearance and disappearance of these species in relation to nucleotides in the medium. Incubation of SR vesicles in ATP for 2 min at 30 °C results in an ATP concentration-dependent increase in nucleotide binding which was resistant to treatment with anion-exchange resin and activated charcoal and inaccessible to enzymes in the medium. The process is saturable at 5 nmol ATP + ADP/mg sarcoplasmic reticulum protein and half-maximal binding occurred with 5 m m ATP. The time dependence of increase of binding showed an initial rapid phase ( t 1 2 ∼- 0.10 min ), followed by a slower phase ( t 1 2 ∼-9 min ). The rapid phase of binding was Ca 2+-dependent with [Ca 2+] 0.5 = 3.5 × 10 −7 m. The slow phase was Ca 2+ independent. An Arrhenius plot of the temperature dependence of the rapid phase of binding showed a break at 24 °C with activation energies of 12.0 and 4.9 kcal mol −1 above and below this point, respectively. The data are consistent with the postulate that the rapid phase of binding of ATP, resulting in a medium-inaccessible species, represents tight binding of ATP to the (Ca 2+, Mg 2+)-ATPase, while the slow nonsaturable process represents leakage into the intravesicular space.

Pulmonary absorption of amino acids in the rat: evidence of carrier transport.

To investigate the absorption of nonmetabolized amino acids from the rat lung, 0.1 ml of Krebs-Ringer phosphate solution containing either 14C-labeled 1-aminocyclopentanecarboxylic acid (cycloleucine) or alpha-aminoisobutyric acid (AIB) was administered to anesthetized animals by way of a catheter introduced through a tight-fitting tracheal cannula. After various times, lungs were removed and assayed for the amount of amino acid that remained. Cycloleucine appeared to be absorbed by a combination of at least two processes: saturable carrier-type transport and a nonsaturable process. The saturable process was inhibited to varying degrees by a number of L- and D-amino acids but not by D-ornithine, betaine, 3-O-methyl-D-glucose, phenol red, disodium cromoglycate, or tetraethylammonium. The nonsaturable process appeared to be too rapid to be accounted for by diffusion alone, thus suggesting interaction with a second carrier system of high capacity. Although AIB was a weak inhibitor of cycloleucine transport, it appeared to be absorbed solely by a nonsaturable process at a rate consistent with diffusion through aqueous membrane channels.

Kinetic characteristics of calcium absorption and secretion by rat colon.

The kinetic characteristics of calcium active transport in rat descending colon were determined by measuring unidirectional transmural calcium fluxes in vitro. The absorptive flux from mucosa to serosa (Jm leads to s) was saturable, with a calculated affinity (Kt) of calcium for the transport system of 1.6 mM and a maximal transport capacity (Vmax) of 133 nmol.cm-2.h-1. The administration of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] increased Jm leads to s by increasing Vmax to 236 nmol.cm-2.h-1 without changing Kt (1.8 mM). The secretory flux from serosa to mucosa (Js leads to m) was not saturable and was not increased by 1,25(OH)2D3. Mannitol, a marker of transepithelial extracellular flux, underwent net absorption in the absence of electrochemical gradients, and its Jm leads to s and Js leads to m were not altered by 1,25(OH)2D3 administration. Addition of 11 mM D-glucose to the bathing medium consistently increased calcium Js leads to m and mannitol Jm leads to s and Js leads to m. Glucose reduced net calcium absorption except when sodium was removed from the medium. Calcium Js leads to m varied linearly with mannitol Js leads to m over the range of medium calcium from 0.125 to 5.0 mM. The behavior of calcium absorption by descending colon is compatible with a carrier-mediated, active-transport mechanism, whereas calcium secretion occurs by a nonsaturable process via a predominately paracellular pathway.

A modified assay method for determining serotonin uptake in human platelets

Effects of various experimental conditions on serotonin (5-HT) uptake in human platelets were examined. The experimental design allowed the evaluation of the effect of diffusion and other non-saturable processes on the affinity and maximum activity of the membrane pump for 5-HT uptake. Total 5-HT uptake was determined by incubating platelet-rich plasma (PRP) with increasing concentrations of serotonin at 37°C for 4 min. The passive uptake was measured by the addition of various 5-HT concentrations to PRP in buffer at 37°C, followed by immediate transfer to an ice-cold water bath. The difference between the total and passive uptake was linear for 6 min. The affinity ( K m) for active platelet serotonin uptake was 0.45 ± 0.09 γmol/ l and maximal rate of uptake ( V) was 10.7 ± 2.1 pmol/10 7 platelets/min. The described method provides a convenient and reliable measure of active 5-HT uptake suitable for clinical investigation. The effect of passive diffusion on kinetic parameters is discussed.

Effect of Ca intake on saturable and nonsaturable components of duodenal Ca transport.

Calcium absorption was studied by an in situ ligated-loop procedure in 9-wk-old male Wistar rats that had been placed from weaning on one of three semisynthetic regimens, 0.17% Ca, 0.44% Ca, or 0.44% Ca plus lactose. Lactose was added because it is known to increase intestinal calcium retention. When the amount of calcium absorbed was expressed as a function of calcium instilled in the loop, it became possible to describe absorption as the sum of a hyperbolic and a linear function, equivalent to a saturable and a nonsaturable process, respectively. The slope of the nonsaturable component was independent of prior calcium intake, while the maximum saturable flux (Jmax) decreased as calcium intake increased. Analysis of the duodenal content of the vitamin D-dependent calcium-binding protein (CaBP, Mr congruent to 10(4)) revealed a positive relation between Jmax and CaBP. Thus, vitamin D appears to be implicated in the saturable, but not in the nonsaturable, component of calcium absorption.

Malondialdehyde alteration of low density lipoproteins leads to cholesteryl ester accumulation in human monocyte-macrophages.

Glutaraldehyde treatment of (125)I-labeled low density lipoprotein ((125)I-native-LDL) produced a modified LDL ((125)I-glut-LDL) with a molecular weight of 10 x 10(6) or more. Malondialdehyde treatment of (125)I-native-LDL produced a product ((125)I-MDA-LDL) with a molecular weight not appreciably different from that of the original lipoprotein. However, the electrophoretic mobility of MDA-LDL indicated a more negative charge than native-LDL. (125)I-MDA-LDL was degraded by two processes: a high-affinity saturable process with maximal velocity at 10-15 mug of protein per ml and a slower, nonsaturable process. The degradation of (125)I-MDA-LDL was readily inhibited by increasing concentrations of nonradioactive MDA-LDL but was not inhibited by acetylated LDL or native-LDL even at concentrations as high as 1600 mug of protein per ml. After exposure of native-LDL to blood platelet aggregation and release in vitro, 1.73 +/- 0.19 nmol of malondialdehyde per mg of LDL protein was bound to the platelet-modified-LDL. No detectable malondialdehyde was recovered from native-LDL that had been treated identically except that the platelets were omitted from the reaction mixture. After incubation with glut-LDL, MDA-LDL, or platelet-modified-LDL for 3 days, human monocyte-macrophages showed a dramatic increase in cholesteryl ester content whereas the cholesteryl ester content of cells incubated with the same concentration of native-LDL did not. Based on these experiments we propose that modification of native-LDL may be a prerequisite to the accumulation of cholesteryl esters within the cells of the atherosclerotic reaction. We further hypothesize that one modification of LDL in vivo may result from malondialdehyde which is released from blood platelets or is produced by lipid peroxidation at the site of arterial injury.

Chlorphentermine uptake by isolated perfused rabbit lung

Studies have shown that many amines accumulate in the lung both in vivo and in vitro. Chlorphentermine (CP) is a phenylethanolamine type of anorexigenic agent in use, known to be accumulated in the lung and to cause pulmonary phospholipidosis. The present study was undertaken to characterize the uptake and accumulation processes of [ 14C]-CP in the lung. These processes were examined in artificially ventilated, recirculating, isolated perfused rabbit lung preparations using either autologous blood or artificial perfusate at initial CP concentrations of 0.125 to 2.0 m m. Perfusate and lung homogenate samples were analyzed for the parent compound and possible metabolite(s). No metabolism of CP was observed. Steady state uptake was reached after 25 min of perfusion. Higher pulmonary extraction of CP was observed from artificial medium than from whole blood. Lung uptake was concentration dependent and was saturable at a concentration of 1.0 m m CP in the perfusate. Higher concentrations resulted in edematous and deteriorating preparations. Total CP uptake by the lungs could be represented by a combination of a non-linear saturable and a linear non-saturable process. Experiments with partial or complete depletion of Na + from the perfusate, inclusion of harmaline (10 −3 and 10 −4 m) or ouabain (10 −3 m), inhibitors of Na +-dependent processes, suggest that CP uptake is partially Na + dependent. In vitro lung slice experiments with Na +-free medium and medium containing harmaline (1 m m) or iodoacetate (1 m m) yielded similar results. Preloading the lung with an equivalent concentration of imipramine, which is known to be concentrated in the lung, resulted in partial inhibition of CP uptake. However, preloading with CP was without effect on imipramine uptake. These results are consistent with a Na +-dependent, carrier-mediated transport process for the uptake of CP by the lung tissue.

Specific interaction of corticosteroids with components of the cell membrane which are involved in the translocation of the hormone into the intravesicular space of purified rat liver plasma membrane vesicles

The uptake of [ 3H]-corticosterone was studied by a highly purified rat liver plasma membrane vesicle fraction. At 23°C, [ 3H]-corticosterone is taken up very rapidly; equilibrium is reached as early as 5 s. At high concentrations of corticosterone (30–8100 nM) uptake occurs predominantly by a non-saturable process, but the presence of a saturable process is also detectable. When the intravesicular space is successively decreased by increasing the osmolarity of the external medium, uptake of [ 3H]-corticosterone decreased. About 45% of the total steroid taken up is translocated into the lumen of the vesicle. In the presence of a 200-fold molar excess of non-labeled corticosterone about 40% of the total uptake of [ 3H]-corticosterone is displaceable. At low concentrations of corticosterone (2—230 nM) and considering only the specific portion of uptake the presence of a high and a low affinity uptake system is observable. The K D of the two systems were 7.2 ± 2.0 nM and 234 ± 67 nM, respectively. The K D of the high affinity system corresponds to the concentration of free corticosterone (about 8 nM) in the plasma of the female rat. The maximal binding capacity was about 180 fmol/(mg membrane protein). Structural analogues of corticosterone reduce the displaceable portion of the uptake of [ 3H]corticosterone; cortisol, progesterone. dexamethasone and 5α-dihydrocorticosterone are strong inhibitors. Among the sex hormones 5α-dihydrotestosterone and diethylstilbestrol are the most efficient inhibitors. These results suggest that corticosterone reacts in a specific manner with components of the plasma membrane which may function to translocate the steroid into the cell.

Uptake of the trypanocidal drug suramin by bloodstream forms of Trypanosoma brucei and its effect on respiration and growth rate in vivo.

After a single intravenous injection of suramin the rate of removal of the drug from the plasma into other tissue compartments of the rat is independent of initial concentration. The data can be fitted to the sum of two exponential functions, consistent with a two-compartment, open model system. Trypanosomes take up only small amounts of suramin in vivo and do not actively concentrate the drug within the cell. Uptake is apparently by a non-saturable process that decreases with time and is dependent on the amount of suramin already taken up. Once within the cell, suramin progressively inhibits respiration and glycolysis, such that, for a given exposure in vivo, inhibition of oxygen consumption is proportional to the total amount of suramin absorbed. It can be calculated that only a fraction (4--9%) of this total is required to inhibit respiration to the extent found in broken cell preparations. The combined inhibition of two key enzymes in glycolysis--the sn-glycerol-3-phosphate oxidase (EC unassigned) and the glycerol-3-phosphate dehydrogenase (NAD+) (sn-glycerol-3-phosphate: NAD+ 2-oxidoreductase, EC 1.1.1.8)--are sufficient to account for the differential inhibition of glucose and oxygen consumption and of pyruvate production, together with the small, but significant, production of glycerol. Even at the highest dose of suramin tolerated by the rat, trypanosomes continue to increase exponentially in the bloodstream for at least 6 h. The mean doubling time is increased from 4.6 h to a maximum of about 12.5 h in rats treated with doses of suramin in the range 25--150 mg/kg. In the light of these and other findings, it is concluded that part of the trypanocidal action of suramin results from the inhibition of ATP production by glycolysis.