Adenosine has several functions within the CNS that involve an inhibitory tone of neurotransmission and neuroprotective actions in pathological conditions. The understanding of adenosine production and release in the brain is therefore of fundamental importance and has been extensively studied. Conflicting results are often obtained regarding the cellular source of adenosine, the stimulus that induces release and the mechanism for release, in relation to different experimental approaches used to study adenosine production and release. A neuronal origin of adenosine has been demonstrated through electrophysiological approaches showing that neurones can release significant quantities of adenosine, sufficient to activate adenosine receptors and to modulate synaptic functions. Specific actions of adenosine are mediated by different receptor subtypes (A(1), A(2A), A(2B) and A(3)), which are activated by various ranges of adenosine concentrations. Another important issue is the measurement of adenosine concentrations in the extracellular fluid under different conditions in order to know the degree of receptor stimulation and understand adenosine central actions. For this purpose, several experimental approaches have been used both in vivo and in vitro, which provide an estimation of basal adenosine levels in the range of 50-200 nM. The purpose of this review is to describe pathways of adenosine production and metabolism, and to summarize characteristics of adenosine release in the brain in response to different stimuli. Finally, studies performed to evaluate adenosine concentrations under physiological and hypoxic/ischemic conditions will be described to evaluate the degree of adenosine receptor activation.

https://onlinelibrary.wiley.com/doi/pdf/10.1046/j.1471-4159.2001.00607.x

Adenosine in the central nervous system: release mechanisms and extracellular concentrations.

Biochemistry, localization and functional roles of ecto-nucleotidases in the nervous system

Adenosine and the concept of ‘retaliatory metabolites’

Publisher Summary Adenosine exerts physiological effects on the nervous system through interactions with cell surface receptors In the nervous system adenosine has been shown to alter levels of cyclic adenosine 3', 5’-monophosphate (CAMP), inhibit synaptic activity and neuronal excitability, and modulate cerebral blood flow, and these effects have led to numerous proposed functional roles for adenosine Adenosine can act as a “retaliatory metabolite” by which cells communicate their energy status to the surrounding tissues Adenosine has been shown to be released during cerebral ischemia, and it increases cerebral blood, inhibits synaptic activity, and decreases neuronal excitability Thus adenosine acts as a neuroprotectant, preventing damage from ischemia and excitotoxicity Adenosine receptors are located throughout the nervous system and extracellular adenosine can effectively modulate neuronal activity This chapter summarizes what is known about how adenosine exerts it effects on neuronal activity, how cells form adenosine and regulate adenosine concentrations, and how cells release adenosine into the extracellular space where it can exert its modulator actions By integrating these different areas of research, it is hoped that a more complete picture of how the brain uses adenosine as a modulator of synaptic activity will be formed The chapter discusses the various adenosine receptors, adenosine formation by S-adenosylhomocysteine hydrolase, adenosine formation metabolism of lntracellular nucleotides, degradation of adenosine, adenosine levels in the brain, extracellular degradation of nucleotides, direct release of adenosine into the extracellular space, and localized adenosine release and synaptic modulation The experimental evidence has made it clear that adenosine serves multiple functions throughout the nervous system Studies on the actions of exogenous adenosine have conclusively shown that adenosine can act as a potent and effective modulator of synaptic activity and neuronal function The purpose of the chapter to summarize what is known about how cells synthesize, release, and regulate adenosine

Role of Adenosine as a Modulator of Synaptic Activity in the Central Nervous System

The subcellular distribution of the ectoenzyme, 5′-nucleotidase, in cerebral cortex and cerebellum of the rat was studied both biochemically and cytochemically. The fractions were characterized biochemically by marker enzymes. The localization of 5′-nucleotidase activity was also investigated cytochemically in the myelin, synaptosomal, mitochondrial, and microsomal fractions. Biochemically 5′-nucleotidase was found to be enriched in the membrane-containing fractions, i.e., myelin, synaptosomal, and microsomal fractions. Cytochemistry showed the reaction product in the myelin fraction to be associated with myelin profiles. In the synaptosomal fraction reaction product could occasionally be seen at synaptosomal membranes, although it could not be attributed unequivocally to the synaptosome itself, since in positions with reaction product unidentifiable membrane structures could always be seen attached. Mitochondria were virtually without any reaction product. In the microsomal fraction 5′-nucleotidase activity was associated with unidentifiable membrane structures. It is concluded that 5′-nucleotidase is associated with myelin profiles and that the high activity found in the synaptosomal fraction is probably not associated with nerve ending plasma membranes.

Subcellular localization of 5'-nucleotidase in rat brain.

The 5'-nucleotidase located in the cytoplasmic fraction of bovine brain cortex was purified to electrophoretic homogeneity. The molecular weight was 134,000 daltons in the presence of sodium deoxycholate, whereas the enzyme formed high molecular weight aggregates in the absence of detergent. The purified enzyme showed the same kinetic and electrophoretic behaviour as the enzyme present in the original cytoplasmic fraction, and the presence of surfactants did not change the Km and Vm values. The nucleotidase from this source was a phosphohydrolase of 5'-mononucleotides acting on the deoxyribonucleotides and ribonucleotides of purines and pyrimidines. 5'-IMP was the preferred substrate; the optimum pH was 7.5. The study of the influence of the temperature on the initial reaction rates allowed calculation of the delta Ea and delta H degrees values. The variation of Vm and Km with a change in pH suggests the existence of a sulfhydryl group and an imidazole group in the enzyme-substrate complex.

Josefa Mallol Montero

Papers

Purification and characterization of bovine brain 5'-nucleotidase.