Effects of iodotubercidin on adenosine kinase activity and nucleoside transport in DDT 1 MF-2 smooth muscle cells

Fiona E. Parkinson, Jonathan D. Geiger

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Abstract

Iodotubercidin is an adenosine kinase inhibitor that through its ability to increase levels of endogenous adenosine can enhance adenosine's receptor- mediated effects. We investigated whether iodotubercidin can inhibit [ 3H]adenosine accumulation by inhibiting transport processes in addition to inhibition of intracellular trapping of labeled adenine nucleotides. Under conditions in which extensive metabolism of intracellular adenosine was present, [ 3H]adenosine accumulation by DDT 1 MF-2 cells was almost completely inhibited by iodotubercidin and the adenosine deaminase inhibitor erythro-9-(2-hydroxy-3-nonyl)-adenine or by the nucleoside transport inhibitor nitrobenzyl-thioinosine. By using similar conditions, [ 3H]adenosine accumulation was significantly greater in Na + buffer than in buffer containing N-methyl-D-glucamine in place of Na +; however, this effect may be explained by an observed 40% inhibition of adenosine kinase activity by N-methyl-D-glucamine. By using uptake intervals of 14 sec to represent the transport component of uptake, iodotubercidin decreased the affinity for adenosine, by about 3-fold, but had no effect on maximum velocity of transport. That these effects of iodotubercidin were due to direct interactions with nucleoside transporters was supported by findings that iodotubercidin inhibited [ 3H]nitrobenzylthioinosine binding to nucleoside transporters with a K(i) value of 4 μM and inhibited [ 3H]uridine and [ 3H]formycin B uptake with IC 50 values of 7 and 15 μM, respectively. These data suggest that iodotubercidin, at pharmacologically relevant concentrations, inhibits nucleoside transport independently of its well characterized inhibition of adenosine kinase and that N-methyl-D-glucamine must be used with caution in experiments to determine the possible presence of Na + gradient-dependent concentrative nucleoside transporters.

Original languageEnglish (US)
Pages (from-to)1397-1401
Number of pages5
JournalJournal of Pharmacology and Experimental Therapeutics
Volume277
Issue number3
StatePublished - Jun 1 1996

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Adenosine Kinase
DDT
Nucleosides
Adenosine
Smooth Muscle Myocytes
Nucleoside Transport Proteins
Thioinosine
Buffers
Adenosine Deaminase Inhibitors
Purinergic P1 Receptors
Adenine Nucleotides
Uridine

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmacology

Cite this

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abstract = "Iodotubercidin is an adenosine kinase inhibitor that through its ability to increase levels of endogenous adenosine can enhance adenosine's receptor- mediated effects. We investigated whether iodotubercidin can inhibit [ 3H]adenosine accumulation by inhibiting transport processes in addition to inhibition of intracellular trapping of labeled adenine nucleotides. Under conditions in which extensive metabolism of intracellular adenosine was present, [ 3H]adenosine accumulation by DDT 1 MF-2 cells was almost completely inhibited by iodotubercidin and the adenosine deaminase inhibitor erythro-9-(2-hydroxy-3-nonyl)-adenine or by the nucleoside transport inhibitor nitrobenzyl-thioinosine. By using similar conditions, [ 3H]adenosine accumulation was significantly greater in Na + buffer than in buffer containing N-methyl-D-glucamine in place of Na +; however, this effect may be explained by an observed 40{\%} inhibition of adenosine kinase activity by N-methyl-D-glucamine. By using uptake intervals of 14 sec to represent the transport component of uptake, iodotubercidin decreased the affinity for adenosine, by about 3-fold, but had no effect on maximum velocity of transport. That these effects of iodotubercidin were due to direct interactions with nucleoside transporters was supported by findings that iodotubercidin inhibited [ 3H]nitrobenzylthioinosine binding to nucleoside transporters with a K(i) value of 4 μM and inhibited [ 3H]uridine and [ 3H]formycin B uptake with IC 50 values of 7 and 15 μM, respectively. These data suggest that iodotubercidin, at pharmacologically relevant concentrations, inhibits nucleoside transport independently of its well characterized inhibition of adenosine kinase and that N-methyl-D-glucamine must be used with caution in experiments to determine the possible presence of Na + gradient-dependent concentrative nucleoside transporters.",
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T1 - Effects of iodotubercidin on adenosine kinase activity and nucleoside transport in DDT 1 MF-2 smooth muscle cells

AU - Parkinson, Fiona E.

AU - Geiger, Jonathan D.

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N2 - Iodotubercidin is an adenosine kinase inhibitor that through its ability to increase levels of endogenous adenosine can enhance adenosine's receptor- mediated effects. We investigated whether iodotubercidin can inhibit [ 3H]adenosine accumulation by inhibiting transport processes in addition to inhibition of intracellular trapping of labeled adenine nucleotides. Under conditions in which extensive metabolism of intracellular adenosine was present, [ 3H]adenosine accumulation by DDT 1 MF-2 cells was almost completely inhibited by iodotubercidin and the adenosine deaminase inhibitor erythro-9-(2-hydroxy-3-nonyl)-adenine or by the nucleoside transport inhibitor nitrobenzyl-thioinosine. By using similar conditions, [ 3H]adenosine accumulation was significantly greater in Na + buffer than in buffer containing N-methyl-D-glucamine in place of Na +; however, this effect may be explained by an observed 40% inhibition of adenosine kinase activity by N-methyl-D-glucamine. By using uptake intervals of 14 sec to represent the transport component of uptake, iodotubercidin decreased the affinity for adenosine, by about 3-fold, but had no effect on maximum velocity of transport. That these effects of iodotubercidin were due to direct interactions with nucleoside transporters was supported by findings that iodotubercidin inhibited [ 3H]nitrobenzylthioinosine binding to nucleoside transporters with a K(i) value of 4 μM and inhibited [ 3H]uridine and [ 3H]formycin B uptake with IC 50 values of 7 and 15 μM, respectively. These data suggest that iodotubercidin, at pharmacologically relevant concentrations, inhibits nucleoside transport independently of its well characterized inhibition of adenosine kinase and that N-methyl-D-glucamine must be used with caution in experiments to determine the possible presence of Na + gradient-dependent concentrative nucleoside transporters.

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