Production of Adenosine from Extracellular ATP at the Striatal Cholinergic Synapse

Abstract

The components of the ectonucleotidase pathway at the immunoaffinity-purified striatal cholinergic synapse have been studied. The ecto-ATPase (EC 3.6.1.15) had a Km of 131 microM, whereas the ecto-ADPase (EC 3.6.1.6) had a Km of 58 microM, was Ca(2+)-dependent, and was inhibited by the ATP analogue 5'-adenylylimidodiphosphate (AMPPNP). The ecto-5'-nucleotidase (EC 3.1.3.5) had a Km of 21 microM, was inhibited by AMPPNP and alpha,beta-methylene ADP, and by a specific antiserum. The Vmax values of the ATPase, ADPase, and 5'-nucleotidase enzymes present at this synapse were in a ratio of 30:14:1. Very little ecto-adenylate kinase activity was detected on these purified synapses. The intraterminal 5'-nucleotidase enzyme, which amounted to 40% of the total 5'-nucleotidase activity, was inhibited by AMPPNP, alpha,beta-methylene ADP, and the antiserum, and also had the same kinetic properties as the ectoenzyme. The time course of ATP degradation to adenosine outside the nerve terminals showed a delay, followed by a period of sustained adenosine production. The delay in adenosine production was proportional to the initial ATP concentration, was a consequence of feedforward inhibition of the ADPase and 5'-nucleotidase, and was inversely proportional to the ecto-5'-nucleotidase activity. The function and characteristics of this pathway and the central role of 5'-nucleotidase in the regulation of extraterminal adenosine concentrations are discussed.

P2 Receptor Excitation of Rodent Hypoglossal Motoneuron ActivityIn VitroandIn Vivo: A Molecular Physiological Analysis

“…The secondary depression may result from diffusion of ATP and activation of nearby inhibitory interneurons (Umemiya and Berger, 1995). However, it is more consistent with hydrolysis of ATP to adenosine and P1(A 1 ) receptor inhibition of glutamatergic activity, as seen in the substantia gelatinosa (Li and Perl, 1995) and striatum (James and Richardson, 1993).…”

Cellular function and molecular structure of ecto-nucleotidases

“…These nucleotides apparently still bind to the catalytic site but without being hydrolyzed [209,210]. Cellular release of ATP/ADP would thus result in feed-forward inhibition of eN and delay of extracellular adenosine formation until their extracellular levels have been reduced to low micromolar levels by other ecto-nucleotidases [211]. Additional functions of eN include the provision of adenosine for cellular reuptake and purine salvage [5] and, as lined out below, potential roles in specific protein (cell) interactions and downstream cellular signaling.…”

5'-Nucleotidases, Nucleosides and their Distribution in the Brain: Pathological and Therapeutic Implications

“…Thus, a small decrease in NTPs levels and/or changes in enzymatic activity of the nucleotide/nucleoside metabolic systems may cause a dramatic alteration in the nucleoside levels [20, [250][251][252][253][254][255]. Extracellular nucleoside triphosphates (such as ATP and UTP) competitively inhibit the activity of e5'NTs, causing a delay in the onset of nucleoside production [58,118,[256][257][258]. Thus, the increased extracellular nucleotide levels (e.g., ATP) in the synaptic cleft (e.g., under hypoxic/ischemic conditions) may cause an accumulation of NMPs (e.g., AMP) to a greater extent compared to nucleosides (e.g., Ado).…”

“…Thus, the increased extracellular nucleotide levels (e.g., ATP) in the synaptic cleft (e.g., under hypoxic/ischemic conditions) may cause an accumulation of NMPs (e.g., AMP) to a greater extent compared to nucleosides (e.g., Ado). The decrease in nucleoside triphosphate levels may result in relief of e5'NT inhibition and thereby elevate nucleoside levels and nucleoside transport into the brain cells anabolized into nucleoside mono-, di-and triphosphates derivatives [58,84,85,257]. During normoxic conditions (at 3.6 mM ATP levels) [133], extracellular nucleotides (e.g., ATP) are mainly catabolized into nucleosides (e.g., Ado) since the level of e5'NT inhibitor nucleoside triphosphates are lower than under ischemic conditions.…”