Identification of lactate as a driving force for prostanoid transport by prostaglandin transporter PGT

Brenda S. Chan1, Shinichi Endo2, Naoaki Kanai3, and Victor L. Schuster1

1 Department of Medicine, Albert Einstein College of Medicine, Bronx, New York 10461; 2 Department of Urology, Yamanashi Medical University, Yamanashi 409-3898; and 3 Department of BioMedical Engineering, Tokai University, Kanagawa 259-11, Japan


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

We previously characterized the prostaglandin (PG) transporter PGT as an exchanger in which [3H]PGE2 influx is coupled to the efflux of a countersubstrate. Here, we cultured HeLa cells that stably expressed human PGT under conditions known to favor glycolysis (glucose as a carbon source) or oxidative phosphorylation (glutamine as a carbon source) and studied the effect on PGT-mediated [3H]PGE2 influx. PGT-expressing cells grown in glutamine exhibited a 2- to 4-fold increase in [3H]PGE2 influx compared with the antisense control, whereas cells grown in glucose exhibited a 14-fold increase. In the presence of 10 vs. 25 mM glucose during the uptake, there was a dose-dependent increment in [3H]PGE2 influx. Cis inhibition of [3H]PGE2 influx occurred with lactate at physiological concentrations (apparent Km = 48 ± 12 mM). Preloading with lactate caused a dose-dependent trans stimulation of PGT-mediated [3H]PGE2 uptake, and external lactate caused trans stimulation of PGT-mediated [3H]PGE2 release. Together, these data are consistent with PGT-mediated PG-lactate exchange. Cells engaged in glycolysis would then be poised energetically for prostanoid uptake by means of PGT.

biological transport; organic anion transport; glycolysis


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

PROSTAGLANDINS (PGS) AND THROMBOXANES have widespread physiological and pathophysiological effects on nearly all cellular processes, including, but not limited to, cardiovascular, gastrointestinal, respiratory, reproductive, renal, and immune systems (56). Because of their broad biological effects, PGs modulate their activity in an autocrine fashion, i.e., they are synthesized by intracellular enzymes at or near their sites of action, exit the cell by simple diffusion (13), and are presented to adjacent PG receptors. Thereafter, extracellular PGs must be metabolized in situ within seconds before they are able to reach the general circulation (17, 21). This loss of biological activity is accomplished through cellular uptake followed by intracellular oxidation (1, 17, 43, 49).

Our laboratory previously identified the novel, broadly expressed PG transporter PGT, whose substrates include PGE2, PGF2alpha , PGD2, and thromboxane-B2 (24). Understanding the molecular mechanism and driving force of PG transport by PGT is necessary to identify its role in PG homeostasis for the organism. PG transport by PGT is Na+, Cl-, and H+ independent and appears to occur by obligate anion exchange (13, 24).

Interestingly, in previous studies from our laboratory, concentrative PG uptake was reduced by glycolysis inhibitors and varied with cellular ATP production (13). There is known coupling between glycolysis and PG metabolism. Inhibition of renal papillary glycolysis by various maneuvers increases net PG release, whereas increasing the supply of glucose decreases PG release (22, 57). Our studies showed that PG transport by PGT exhibited a time-dependent overshoot that resulted in transient accumulation of [3H]PGE2 (13), which is consistent with exchange in which [3H]PGE2 influx is coupled to the efflux of a countersubstrate that exhibits a falling outwardly directed gradient. Taken together, the data suggested that PGT-mediated influx of PGs may be coupled to the efflux of an intracellular metabolic end product involved in the generation of ATP by glycolysis.

In the present study, we cultured HeLa cells that stably express PGT under conditions known to favor glycolysis or oxidative phosphorylation and studied the effect on PGT-mediated transport. Our data indicate that the direction and magnitude of PG transport by means of PGT vary directly with the transmembrane concentration gradient of lactate, which is consistent with PG-lactate exchange.


    MATERIALS AND METHODS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Materials

Glucose-6-phosphate, fructose-6-phosphate, fructose-1,6-diphosphate, glyceraldehyde-3-phosphate, 1,3,-diphosphoglycerate, 3-phosphoglycerate, 2-phosphoglycerate, phosphoenolpyruvate, pyruvate, and lactate were purchased from Sigma and were the purest grade available. [3H]PGE2 was from DuPont-New England Nuclear.

Stable Transfection of PGT in HeLa Cells

HeLa cells (ATCC) were grown in Dulbecco's modified Eagle's medium plus 5% fetal bovine serum and 100 U/ml penicillin-streptomycin (GIBCO). Full-length sense and antisense PGT cDNAs were cloned into the vector pMEP4 (Invitrogen). HeLa cells were transfected with Lipofectin (GIBCO) and pMEP4-antisense human PGT or pMEP4-sense human PGT and cultured in selective media containing hygromycin (600 ug/ml; GIBCO) 48 h later. After 14-18 days, resistant colonies were selected and expanded in selective media.

Cell Culture

Stable transfectants were maintained in humidified incubators with 5% CO2 at 37°C in Dulbecco's modified Eagle's medium plus 5% fetal bovine serum and 100 U/ml penicillin-streptomycin. In experiments analyzing the effects of glycolysis on PGT-mediated transport, stable transfectants were grown overnight as monolayers on 35-mm dishes in Dulbecco's modified Eagle's medium with 2 mM glutamine, glutamine plus 25 mM glucose, or glutamine plus 25 mM 2-deoxyglucose supplemented with 5% fetal bovine serum.

Transport Assays

Influx measurements. The cell monolayers were washed twice with a balanced salt solution [BSS; (in mM) 135 NaCl, 13 H-HEPES, 13 Na-HEPES, 2.5 CaCl2, 1.2 MgCl2, 0.8 MgSO4, and 5 KCl]. Influx measurements were initiated by the addition of [3H]PGE2 to the flux media (BSS). Influx measurements were carried out at room temperature over 10 min and were terminated by aspiration of the incubation media followed by two rapid washings with ice-cold 5% BSA in BSS and two additional washings with ice-cold BSS. Cells were scraped into 1 ml saline, mixed with a liquid scintillation cocktail (National Diagnostics), and analyzed by liquid scintillation counting. Influx values were calculated as femtomoles per milligram protein per nanomolar concentration [3H]PGE2 and expressed as means ± SE from duplicate monolayers. In some experiments, 10-min uptakes were performed in the presence of BSS vs. BSS plus 10 or 25 mM glucose.

Cis-inhibition. In cis-inhibition studies, uptakes were performed in the presence of unlabeled substrates (glucose-6-phosphate, fructose-6-phosphate, fructose-1,6-diphosphate, glyceraldehyde-3-phosphate, 1,3,-diphosphoglycerate, 3-phosphoglycerate, 2-phosphoglycerate, phosphoenolpyruvate, pyruvate, and lactate). Candidate substrates were added at 10 mM concentrations during [3H]PGE2 uptake. The half-maximal inhibition constant (K1/2) was determined by relating the uninhibited uptake rate to the inhibited uptake rate calculated from the relationship
K<SUB>½</SUB>=[V(i)<IT>/</IT>V<IT>−</IT>V(i)]<IT>·K</IT><SUB>m</SUB><IT>·</IT>[i]<IT>/</IT>[S]<IT>+K</IT><SUB>m</SUB>
where V is uptake without inhibitor, V(i) is uptake with inhibitor, [i] is inhibitor concentration, and [S] is substrate concentration (37).

Trans-stimulation. In some experiments, stable transfectants were incubated for 15 min in varying concentrations of lactate, washed twice, and then subjected to the [3H]PGE2 uptake assay.

In other studies, monolayers were preloaded with [3H]PGE2 for 20 min. Immediately thereafter, cells were washed twice with room temperature BSS to remove adherent [3H]PGE2. At time 0, 1 ml of BSS ± 25 mM lactate was added to each monolayer. At each subsequent time interval (2- to 10-min intervals), the 1 ml of efflux media was removed for scintillation counting and 1 ml of fresh BSS with or without 25 mM lactate was added. At the end of the experiment, the cell monolayers were scraped and analyzed by scintillation counting. The remaining counts per minute in the cells were added to the sum of the efflux counts to estimate the amount of isotope that had been initially loaded. Efflux values were calculated as the percentage ± SE of the total [3H]PGE2 released at 2-, 5-, and 10-min intervals.

Metabolites

Lactate and pyruvate concentrations were measured in the extracellular media by using a colorimetric assay on the basis of the enzymatic conversion of lactic acid to pyruvate and hydrogen peroxide by lactate oxidase (Sigma). Pyruvate concentrations were determined by using a colorimetric assay on the basis of the enzymatic conversion of pyruvate and NADH to lactate and NAD by lactate dehydrogenase (Sigma). Intracellular ATP concentrations of monolayers grown in media with or without glucose were determined by using an ATP assay (Sigma) on the basis of the enzymatic reaction described by Bucher (12).


    RESULTS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Our laboratory previously reported that the PG transporter PGT is probably an obligatory, electrogenic anion exchanger (13). Moreover, our laboratory demonstrated that transport is reduced by inhibiting oxidative phosphorylation and glycolysis, suggesting that the countersubstrate(s) may be a product(s) of intermediary metabolism (13). On the basis of these observations, we cultured HeLa cells that stably expressed human PGT under conditions known to favor glycolysis or oxidative phosphorylation, and then we studied the effect on PGT-mediated [3H]PGE2 influx.

HeLa cells stably transfected with pMEP4-antisense human PGT showed virtually no uptake of [3H]PGE2, which is consistent with previous data from our laboratory (13, 33, 45) and others (4, 8) that PGs at physiological pH enter cells poorly using simple diffusion. In contrast, HeLa cells transfected with pMEP4-sense human PGT showed a 27-fold increase in [3H]PGE2 uptake over that in antisense control (data not shown). All subsequent experiments were performed with the stably expressing sense PGT HeLa cells.

It has been well established that cells in culture can utilize either glutamine or glucose to provide cell energy, depending on the availability of the respective substrates (9, 30, 46, 62). To test the hypothesis that PGT-mediated transport is dependent on metabolism, we examined the degree to which transport is dependent on oxidative phosphorylation or glycolysis by incubating our stable transfectants overnight in media containing 2 mM glutamine with or without 25 mM glucose. Changing the pattern of metabolism had no effect on cell morphology; however, the cell-doubling times were ~33 h with glutamine and 24 h with glutamine plus glucose. No significant difference in the intracellular ATP concentrations were noted (data not shown), which is consistent with reports in the literature (46). However, there was a marked difference in the ability of PGT-expressing PGT cells to accumulate [3H]PGE2 as a function of glycolysis. As shown in Fig. 1, PGT-expressing cells grown in glutamine plus 25 mM glucose showed a 14-fold increase in [3H]PGE2 influx compared with PGT-expressing cells grown in glutamine without glucose or in glutamine with 25 mM 2-deoxyglucose. Similarly, [3H]PGE2 influx in the absence or presence of 10 or 25 mM glucose during 10-min transport assays showed a dose-dependent increment (Fig. 2). In contrast, 10-min uptakes in the presence of 10 or 25 mM 2-deoxyglucose, as an osmotic control, were not different from control uptake (Fig. 1). Taken together, the data indicate that the rate of [3H]PGE2 influx appears to be influenced by the rate of glycolysis.


View larger version (13K):
[in this window]
[in a new window]
 
Fig. 1.   Effect of culture media on protaglandin transporter PGT-mediated [3H]PGE2 uptake. HeLa cells stably transfected with sense PGT were grown overnight in media containing 2 mM glutamine with or without 25 mM glucose or 2 mM glutamine with 25 mM 2-deoxyglucose as indicated. Values are means ± SE of 3 experiments.



View larger version (11K):
[in this window]
[in a new window]
 
Fig. 2.   Effect of glucose concentration on PGT-mediated [3H]PGE2 uptake. pMEP4-PGT stable transfectants were exposed to varying concentrations of glucose during 10-min transport assays. Values are percent increase ± SE of 3 experiments compared with 0 glucose.

To test intermediary metabolites as candidate substrates, we performed cis-inhibition experiments by adding metabolites to the outside of the cell during [3H]PGE2 influx. External substrates other than PGE2 would be expected to reorient the exchanger toward the cytoplasm, rendering it unavailable to bind and internalize external tracer PGE2. As shown in Table 1, several glycolytic products inhibited [3H]PGE2 influx; however, only lactate inhibited [3H]PGE2 influx at a concentration that might be approached in the cytoplasm. In accordance with this concept, in cells grown in glutamine overnight, the lactate concentration of the external media (which is likely in equilibrium with the cytoplasm at that point) was 3.5 ± 0.5 vs. 29 ± 6.1 mM in cells grown in glutamine plus glucose. This sevenfold increase in lactate concentration compares favorably with the sixfold increase in [3H]PGE2 uptake in cells grown in the presence of glucose. In contrast, medium pyruvate concentrations were similar in the two groups (119 ± 13 vs. 123 ± 18 µM). We cannot exclude the possibility that under nonglycolytic conditions, metabolites other than the lactate shown in Table 1 might serve as substrates for PGT.

                              
View this table:
[in this window]
[in a new window]
 
Table 1.   Cis-inhibition of [3H]PGE2 uptake by glycolytic end products

If lactate serves as the countersubstrate for PGE2 on the PGT anion exchanger, then the rate of lactate efflux should vary with the rate of PGE2 accumulation. As demonstrated in Fig. 3, net [3H]PGE2 uptake decreased pari passu with the rate of lactate release over time, suggesting that PGT-mediated transport is dependent on intracellular lactate. This is consistent with coupling between [3H]PGE2 uptake and lactate efflux by means of PGT.


View larger version (14K):
[in this window]
[in a new window]
 
Fig. 3.   Relationship of time-dependent PGT-mediated net [3H]PGE2 accumulation and lactate efflux. Ability of PGT-expressing cells to accumulate [3H]PGE2 decreases as lactate efflux decreases over time. Values are means ± SE of 2 experiments.

If lactate is transported by PGT, it should trans accelerate PGE2 transport, i.e., cytoplasmic lactate would be expected to reorient the exchanger toward the extracellular space, rendering it more available to bind and internalize external tracer PGE2. To test this, we incubated the stable transfectants overnight in glutamine media so that the intracellular lactate concentrations before the experiment were low. Then, we incubated the cells in BSS in the absence or presence of 10 or 25 mM lactate for 15 min before the [3H]PGE2 transport assay. As shown in Fig. 4, this produced a dose-dependent increase in [3H]PGE2 uptake, i.e., lactate trans stimulated [3H]PGE2 uptake.


View larger version (11K):
[in this window]
[in a new window]
 
Fig. 4.   Effect of lactate preincubation on subsequent PGT-mediated net [3H]PGE2 uptake. Stable transfectants grown overnight in glutamine media were preincubated with balanced salt solution (BSS) or BSS plus 10 or 25 mM lactate for 15 min before [3H]PGE2 transport assays. Values are percent increase ± SE of 5 experiments compared with 0 lactate.

The converse experiments were also performed. Stable transfectants were loaded with [3H]PGE2, and the rate of [3H]PGE2 efflux was measured in the absence and presence of extracellular lactate. As shown in Table 2, trans lactate caused a significant increase in [3H]PGE2 efflux, consistent with PGT-mediated lactate-PGE2 anion exchange.

                              
View this table:
[in this window]
[in a new window]
 
Table 2.   [3H]PGE2 efflux in cells incubated in balanced salt solution in presence and absence of 25 mM lactate


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

We previously found that PGT-mediated PG uptake varied directly with intracellular ATP concentration (13). However, as PGT demonstrates no ATP binding motifs and has no homology to the P-type ATPases, we concluded that ATP is probably indirectly involved in PG transport. We therefore hypothesized that production of an intracellular countersubstrate coupled to PG uptake may be dependent on cellular metabolism. In the present study, we established HeLa cells that stably expressed human PGT and studied the effect of altering glucose metabolism on PGT-mediated [3H]PGE2 uptake. We favored oxidative phosphorylation or glycolysis by supplying cells with glutamine or glutamine plus glucose, respectively, in an overnight incubation. We found that the presence or absence of glucose in the culture media does not result in significant changes in ATP levels. These results are in accordance with those of Reitzer et al. (46), who also found no change in intracellular citric acid intermediates with these maneuvers. Nonetheless, this maneuver results in extremely variable levels of glycolytic intermediates (46). Our data clearly indicate that substantial PG influx could not be supported in the absence of glycolysis. On the other hand, acute exposure of cells to glucose for only 10 min during the transport assay caused a dose-dependent increase in PGT-mediated PG uptake, which is consistent with the coupling of transport to glucose metabolism.

Several results presented here support intracellular lactate as the driving force for prostanoid uptake by means of PGT. First, the degree of increase in lactate concentration in cells cultured in glucose was similar to the degree of increase in PG uptake. Second, the time-dependent net accumulation of [3H]PGE2 varied directly with the rate of lactate efflux. Third, preloading cells with lactate resulted in a dose-dependent augmentation of subsequent [3H]PGE2 influx. Taken together, these data provide strong evidence that intracellular lactate drives PGT-mediated PG uptake.

We hypothesize that lactate is coupled to PG uptake by means of lactate-PG exchange. Several glycolytic metabolites cis inhibited PG influx; however, the Km for lactate is closer to known cytoplasmic lactate levels than those for the other metabolites (7, 19, 34, 48, 52, 54). Indeed, the medium lactate concentration of cells cultured in the presence and absence of glucose was 29 ± 8.5 vs. 3.5 ± 0.7 mM, respectively. On the assumption that the cytoplasmic and medium lactate concentrations are at equilibrium after overnight incubation, these data mean that the lactate concentration within cells was well within the range to serve as a substrate for PGT. In contrast, pyruvate was present at low levels in cells cultured with and without glucose [medium (pyruvate) = 119 vs. 123 µM, respectively], which argued against it being a substrate. Further evidence for lactate-PG exchange is the trans acceleration of [3H]PGE2 efflux by the presence of external lactate; i.e., an inwardly directed lactate gradient results in lactate influx in exchange for PG efflux.

Lactate production under aerobic conditions has been described in many tissues, including lung (28, 32, 40, 47), kidney (38, 60), brain (36, 42), vascular smooth muscle (5, 35, 41), heart (2, 20), and skeletal muscle (11, 23). The presence of outwardly directed lactate gradients in diverse cell types suggests that PGT is poised for PG uptake rather than for release in vivo. Accordingly, using a number of in vitro expression systems, we have failed to demonstrate augmented PG release from cells expressing PGT; on the other hand, we observed that expression of PGT at the plasma membrane results in less net PG release compared with controls (Chan BS, Bao Y, and Schuster VL, unpublished observations).

Taken together, these results suggest that PGT may be involved in reuptake of newly synthesized PGs. This model is similar to that of synaptic signaling, in which plasma membrane neurotransmitter transporters regulate extracellular neurotransmitter levels by uptake. Both prostanoids and neurotransmitters are synthesized and released at their sites of action, where they activate cell surface receptors. In the case of neurotransmission, termination of signaling is accomplished by local reuptake by high-affinity transporters (25, 26, 61). Here, we propose that net release of PGs is controlled by PGT-mediated reuptake within or near cells that release PGs. In concordance with this hypothesis, recent data from our laboratory have localized PGT to cells known to synthesize and release PGs (3).

A reuptake model can explain two previously puzzling papers that showed that inhibiting glycolysis in renal papillae increases the rate of PGE2 release. Herman et al. (22) found that incubation of renal papillae with 2-deoxyglucose or the glycolytic inhibitor NaF increased net PGE2 release. Similarly, Tannenbaum et al. (57) found that increasing amounts of glucose in the buffer suppressed net PGE2 release. One explanation for these findings is that glycolysis promotes PG reuptake by PGT, resulting in a reduction in net PG release.

This hypothesis has ramifications for control of circulation and has potential for explaining vasomotor control during tissue ischemia. Occlusion of blood flow causes release of PGs and PG-mediated vasodilatation (6, 10, 15, 58). In our model, an increase in extracellular lactate in ischemic tissue would create an adverse gradient for PGT-mediated PG reuptake into cells such that the net release of PGs would be augmented.

In addition, Kawamura et al. (27) showed that the antiapoptotic effect of PGE in PC-12 cells requires PG uptake by means of PGT. This raises the possibility that extracellular PGs are taken up by cells to signal cell proliferation. In the case of some malignancies, cell proliferation involves abrogation of apoptosis (14, 39, 44, 51, 59). Tumor cells generate large amounts of lactic acid under aerobic conditions (18, 29, 55), thereby creating a favorable gradient for PG uptake and any associated antiapoptosis. In addition, smooth muscle cell proliferation, as observed in atherosclerosis, pulmonary hypertension, and venous pathologies, appears to be mediated, in part, by PGD2, PGF2alpha , and PGE (31, 50, 53). This mitogenic response is triggered by exposure of endothelial cells to hypoxia, suggesting that the pro-proliferative effect of PGs may be under metabolic control by means of PGT-mediated uptake into the cell.

In summary, our data demonstrate that PGT-mediated uptake of PGs is dependent on glycolysis and that transport is coupled to lactate by a mechanism consistent with lactate-PG exchange. The present model supports a role for PGT in the uptake of PGs in vivo and provides a molecular explanation for events in which metabolic derangement affects PG release and activity.


    ACKNOWLEDGEMENTS

This work was supported by National Institute of Diabetes and Digestive and Kidney Diseases Grants RO1-DK-49688 and KO8-DK-02492.


    FOOTNOTES

Address for reprint requests and other correspondence: B. S. Chan, Renal Div., Albert Einstein College of Medicine, Ullmann Bldg., Rm. 615, 1300 Morris Park Ave., Bronx, NY 10461.

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

10.1152/ajprenal.00151.2001

Received 15 May 2001; accepted in final form 26 December 2001.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

1.   Anderson, MW, and Eling TE. Prostaglandin removal and metabolism by isolated perfused rat lung. Prostaglandins 11: 645-677, 1976[Medline].

2.   Ascuitto, RJ, Joyce JJ, and Ross-Ascuitto NT. Mechanical function and substrate oxidation in the neonatal pig heart subjected to pacing-induced tachycardia. Mol Genet Metab 66: 212-223, 1999[ISI][Medline].

3.   Bao, Y, Pucci ML, Chan BS, Lu R, Ito S, and Schuster VL. Prostaglandin transporter PGT is expressed in cell types that synthesize and release prostanoids. Am J Physiol Renal Physiol 282: F1103-F1110, 2002[Abstract/Free Full Text].

4.   Baroody, RA, and Bito LZ. The impermeability of the basic cell membrane to thromboxane-B2' prostacyclin and 6-keto-PGF 1 alpha. Prostaglandins 21: 133-142, 1981[Medline].

5.   Barron, JT, and Parrillo JE. Production of lactic acid and energy metabolism in vascular smooth muscle: effect of dichloroacetate. Am J Physiol Heart Circ Physiol 268: H713-H719, 1995[Abstract/Free Full Text].

6.   Beamish, RE, Das PK, Karmazyn M, and Dhalla NS. Prostaglandins and heart disease. Can J Cardiol 1: 66-74, 1985[Medline].

7.   Berkowitz, BA, Bansal N, and Wilson CA. Non-invasive measurement of steady-state vitreous lactate concentration. NMR Biomed 7: 263-268, 1994[ISI][Medline].

8.   Bito, LZ, and Baroody RA. Impermeability of rabbit erythrocytes to prostaglandins. Am J Physiol 229: 1580-1584, 1975[Abstract/Free Full Text].

9.   Brand, RM, Lyons RH, and Midgley AR. Understanding the dynamics of cellular responsiveness to modifications of metabolic substrates in perfusion. J Cell Physiol 160: 10-16, 1994[ISI][Medline].

10.   Brezis, M, Rosen S, Stoff JS, Spokes K, Silva P, and Epstein FH. Inhibition of prostaglandin synthesis in rat kidney perfused with and without erythrocytes: implication for analgesic nephropathy. Miner Electrolyte Metab 12: 326-332, 1986[ISI][Medline].

11.   Brooks, GA. Lactate production under fully aerobic conditions: the lactate shuttle during rest and exercise. Federation Proc 45: 2924-2929, 1986[ISI][Medline].

12.   Bucher, T. Uber ein phosphatubertragendes Garungsferment. Biochim Biophys Acta 1000: 228-250, 1989[Medline].

13.   Chan, BS, Satriano JA, Pucci M, and Schuster VL. Mechanism of prostaglandin E2 transport across the plasma membrane of HeLa cells and Xenopus oocytes expressing the prostaglandin transporter "PGT." J Biol Chem 273: 6689-6697, 1998[Abstract/Free Full Text].

14.   Compagni, A, and Christofori G. Recent advances in research on multistage tumorigenesis. Br J Cancer 83: 1-5, 2000[ISI][Medline].

15.   Dunn, MJ, Scharschmidt L, and Zambraski E. Mechanisms of the nephrotoxicity of non-steroidal anti-inflammatory drugs. Arch Toxicol Suppl 7: 328-337, 1984[Medline].

16.   Feigen, LP, Chapnick BM, Flemming JE, Flemming JM, and Kadowitz PJ. Renal vascular effects of endoperoxide analogs, prostaglandins, and arachidonic acid. Am J Physiol Heart Circ Physiol 233: H573-H579, 1977[ISI][Medline].

17.   Ferreira, SH, and Vane JR. Prostaglandins: their disappearance from and release into the circulation. Nature 216: 868-873, 1967[ISI][Medline].

18.   Floridi, A, Paggi MG, and Fanciulli M. Modulation of glycolysis in neuroepithelial tumors. J Neurosurg Sci 33: 55-64, 1989[Medline].

19.   Gfrerer, RJ, Brunner GA, Trajanoski Z, Schaupp L, Sendlhofer G, Skrabal F, Jobst G, Moser I, Urban G, Pieber TR, and Wach P. Novel system for real-time ex vivo lactate monitoring in human whole blood. Biosens Bioelectron 13: 1271-1278, 1998[ISI][Medline].

20.   Goodwin, GW, Ahmad F, Doenst T, and Taegtmeyer H. Energy provision from glycogen, glucose, and fatty acids on adrenergic stimulation of isolated working rat hearts. Am J Physiol Heart Circ Physiol 274: H1239-H1247, 1998[Abstract/Free Full Text].

21.   Hamberg, M, and Samuelsson B. On the metabolism of prostaglandins E1 and E2 in man. J Biol Chem 246: 6713-6721, 1971[Abstract/Free Full Text].

22.   Herman, CA, Zenser TV, and Davis BB. Prostaglandin E2 production by renal inner medullary tissue slices: effect of metabolic inhibitors. Prostaglandins 14: 679-687, 1977[Medline].

23.   James, JH, Wagner KR, King JK, Leffler RE, Upputuri RK, Balasubramaniam A, Friend LA, Shelly DA, Paul RJ, and Fischer JE. Stimulation of both aerobic glycolysis and Na+-K+-ATPase activity in skeletal muscle by epinephrine or amylin. Am J Physiol Endocrinol Metab 277: E176-E186, 1999[Abstract/Free Full Text].

24.   Kanai, N, Lu R, Satriano J, Bao Y, Wolkoff AW, and Schuster VL. Identification and characterization of a prostaglandin transporter. Science 268: 866-869, 1995[ISI][Medline].

25.   Kanner, BI, and Schuldiner S. Mechanism of transport and storage of neurotransmitters. CRC Crit Rev Biochem 22: 1-38, 1987[ISI][Medline].

26.   Kavanaugh, MP, Arriza JL, North RA, and Amara SG. Electrogenic uptake of gamma-aminobutyric acid by a cloned transporter expressed in Xenopus oocytes. J Biol Chem 267: 22007-22009, 1992[Abstract/Free Full Text].

27.   Kawamura, T, Horie S, Maruyama T, Akira T, Imagawa T, and Nakamura N. Prostaglandin E1 transported into cells blocks the apoptotic signals induced by nerve growth factor deprivation. J Neurochem 72: 1907-1914, 1999[ISI][Medline].

28.   Kerr, JS, Baker NJ, Bassett DJ, and Fisher AB. Effect of perfusate glucose concentration on rat lung glycolysis. Am J Physiol Endocrinol Metab Gastrointest Physiol 236: E229-E233, 1979[Abstract/Free Full Text].

29.   Kristjansen, PE, Brown TJ, Shipley LA, and Jain RK. Intratumor pharmacokinetics, flow resistance, and metabolism during gemcitabine infusion in ex vivo perfused human small cell lung cancer. Clin Cancer Res 2: 359-367, 1996[Abstract].

30.   Lanks, KW. End products of glucose and glutamine metabolism by L929 cells. J Biol Chem 262: 10093-10097, 1987[Abstract/Free Full Text].

31.   Loesberg, C, van Wijk R, Zandbergen J, van Aken WG, van Mourik JA, and de Groot PG. Cell cycle-dependent inhibition of human vascular smooth muscle cell proliferation by prostaglandin E1. Exp Cell Res 160: 117-125, 1985[ISI][Medline].

32.   Longmore, WJ, and Mourning JT. Lactate production in isolated perfused rat lung. Am J Physiol 231: 351-354, 1976[Abstract/Free Full Text].

33.   Lu, R, Kanai N, Bao Y, and Schuster VL. Cloning, in vitro expression, and tissue distribution of a human prostaglandin transporter cDNA (hPGT). J Clin Invest 98: 1142-1149, 1996[Abstract/Free Full Text].

34.   Lukkarainen, J, Kauppinen RA, Koistinaho J, Halmekyto, Alhonen LM, and Janne J. Cerebral energy metabolism and immediate early gene induction following severe incomplete ischaemia in transgenic mice overexpressing the human ornithine decarboxylase gene: evidence that putrescine is not neurotoxic in vivo. Eur J Neurosci 7: 1840-1849, 1995[ISI][Medline].

35.   Lynch, RM, and Paul RJ. Glucose uptake in porcine carotid artery: relation to alterations in active Na+-K+ transport. Am J Physiol Cell Physiol 247: C433-C440, 1984[Abstract].

36.   Magistretti, PJ, and Pellerin L. Cellular mechanisms of brain energy metabolism and their relevance to functional brain imaging. Philos Trans R Soc Lond B Biol Sci 354: 1155-1163, 1999[ISI][Medline].

37.   Neame, KD, and Richards TG. Elementary Kinetics of Membrane Carrier Transport. New York: Wiley, 1972, p. 56-79.

38.   Needleman, P, Passonneau JV, and Lowry OH. Distribution of glucose and related metabolites in rat kidney. Am J Physiol 215: 655-659, 1968[Free Full Text].

39.   Negoescu, A. Apoptosis in cancer: therapeutic implications. Histol Histopathol 15: 281-297, 2000[ISI][Medline].

40.   O'Neil, JJ, and Tierney DF. Rat lung metabolism: glucose utilization by isolated perfused lungs and tissue slices. Am J Physiol 226: 867-873, 1974[Free Full Text].

41.   Paul, RJ, Krisanda JM, and Lynch RM. Vascular smooth muscle energetics. J Cardiovas Pharmacol 6, Suppl2: S320-S327, 1984[ISI][Medline].

42.   Pellerin, L, and Magistretti PJ. Glutamate uptake into astrocytes stimulates aerobic glycolysis: a mechanism coupling neuronal activity to glucose utilization. Proc Natl Acad Sci USA 91: 10625-10629, 1994[Abstract/Free Full Text].

43.   Piper, PJ, Vane JR, and Wyllie JH. Inactivation of prostaglandins by the lungs. Nature 225: 600-604, 1970[ISI][Medline].

44.   Polverini, PJ, and Nor JE. Apoptosis and predisposition to oral cancer. Crit Rev Oral Biol Med 10: 139-152, 1999[Abstract].

45.   Pucci, ML, Bao Y, Chan B, Itoh S, Lu R, Copeland NG, Gilbert DJ, Jenkins NA, and Schuster VL. Cloning of mouse prostaglandin transporter PGT cDNA: species-specific substrate affinities. Am J Physiol Regulatory Integrative Comp Physiol 277: R734-R741, 1999[Abstract/Free Full Text].

46.   Reitzer, LJ, Wice BM, and Kennell D. Evidence that glutamine, not sugar, is the major energy source for cultured HeLa cells. J Biol Chem 254: 2669-2676, 1979[ISI][Medline].

47.   Rhoades, RA, Shaw ME, Eskew ML, and Wali S. Lactate metabolism in perfused rat lung. Am J Physiol Endocrinol Metab Gastrointest Physiol 235: E619-E623, 1978[Abstract/Free Full Text].

48.   Rovetto, MJ, Lamberton WF, and Neely JR. Mechanisms of glycolytic inhibition in ischemic rat hearts. Circ Res 37: 742-751, 1975[Abstract].

49.   Samuelsson, B. Biosynthesis of prostaglandins. Federation Proc 31: 1442-1450, 1972[ISI][Medline].

50.   Schror, K, and Weber AA. Roles of vasodilatory prostaglandins in mitogenesis of vascular smooth muscle cells. Agents Actions Suppl 48: 63-91, 1997[Medline].

51.   Schulte-Hermann, R, Bursch W, Grasl-Kraupp B, Marian B, Torok L, Kahl-Rainer P, and Ellinger A. Concepts of cell death and application to carcinogenesis. Toxicol Pathol 25: 89-93, 1997[ISI][Medline].

52.   Simonsen, L, Holstein P, Larsen K, and Bulow J. Glucose metabolism in chronic diabetic foot ulcers measured in vivo using microdialysis. Clin Physiol 18: 355-359, 1998[ISI][Medline].

53.   Sinzinger, H, Zidek T, Fitscha P, Kaliman J, and Steurer G. Platelet derived growth factor (PDGF) and prostaglandins (PGE1, PGI2) as modulators of the atherogenetic process. Folia Haematol Int Mag Klin Morphol Blutforsch 115: 439-442, 1988[Medline].

54.   Smith, EW, Skelton MS, Kremer DE, Pascoe DD, and Gladden LB. Lactate distribution in the blood during steady-state exercise. Med Sci Sports Exerc 30: 1424-1429, 1998[ISI][Medline].

55.   Smith, TA, and Titley J. Deoxyglucose uptake by a head and neck squamous carcinoma: influence of changes in proliferative fraction. Int J Radiat Oncol Biol Phys 47: 219-223, 2000[ISI][Medline].

56.   Smith, WL. The eicosanoids and their biochemical mechanisms of action. Biochem J 259: 315-324, 1989[ISI][Medline].

57.   Tannenbaum, J, Sweetman BJ, Nies AS, Aulsebrook K, and Oates JA. The effect of glucose on the synthesis of prostaglandins by the renal papilla of the rat in vitro. Prostaglandins 17: 337-350, 1979[Medline].

58.   Weir, EK, McMurtry IF, Tucker A, Reeves JT, and Grover RF. Prostaglandin synthetase inhibitors do not decrease hypoxic pulmonary vasoconstriction. J Appl Physiol 41: 714-718, 1976[Abstract/Free Full Text].

59.   Williams, MA, Newland AC, and Kelsey SM. The potential for monocyte-mediated immunotherapy during infection and malignancy. Part I: apoptosis induction and cytotoxic mechanisms. Leuk Lymphoma 34: 1-23, 1999[ISI][Medline].

60.   Yorita, K, Yamano T, Ikeda K, Kobayashi T, Shiota M, and Sugano T. Distribution of glycolysis and gluconeogenesis in perfused chicken kidney. Am J Physiol Regulatory Integrative Comp Physiol 253: R679-R686, 1987[Abstract/Free Full Text].

61.   Zerangue, N, and Kavanaugh MP. Flux coupling in a neuronal glutamate transporter. Nature 383: 634-637, 1996[ISI][Medline].

62.   Zielke, HR, Ozand PT, Tildon JT, Sevdalian DA, and Cornblath M. Reciprocal regulation of glucose and glutamine utilization by cultured human diploid fibroblasts. J Cell Physiol 95: 41-48, 1978[ISI][Medline].


Am J Physiol Renal Fluid Electrolyte Physiol 282(6):F1097-F1102
0363-6127/02 $5.00 Copyright © 2002 the American Physiological Society