E. Sylvester Vizi

Neurochemistry and pharmacology of the major hippocampal transmitter systems: Synaptic and nonsynaptic interactions

Hippocampus plays a crucial role in important brain functions (e.g. memory, learning) thus in the past two decades this brain region became a major objective of neuroscience research. During this period large number of anatomical, neurochemical and electrophysiological data have been accumulated. While excellent reviews have been published on the anatomy and electrophysiology of hippocampal formation, the neurochemistry of this area has not been thoroughly surveyed. Therefore the aim of this review is to summarize the neurochemical and pharmacological data on the release of the major neurotransmitters found in the hippocampal region: glutamate (GLU), γ‐amino butyric acid (GABA), acetylcholine (ACh), noradrenaline (NA) and serotonin (5‐HT). In addition, this review analyzes the synaptic and nonsynaptic interactions between hippocampal neuronal elements and overviews how auto‐ and heteroreceptors are involved in the presynaptic modulation of transmitter release. The presented data clearly show that transmitters released from axon terminals without synaptic contact play an important role in the fine tuning of communication between neurons within a neuronal circuit. Hippocampus 1998;8:566–607.

CD39 and CD73 in immunity and inflammation

The enzymatic activities of CD39 and CD73 play strategic roles in calibrating the duration, magnitude, and chemical nature of purinergic signals delivered to immune cells through the conversion of ADP/ATP to AMP and AMP to adenosine, respectively. This drives a shift from an ATP-driven proinflammatory environment to an anti-inflammatory milieu induced by adenosine. The CD39/CD73 pathway changes dynamically with the pathophysiological context in which it is embedded. It is becoming increasingly appreciated that altering this catabolic machinery can change the course or dictate the outcome of several pathophysiological events, such as AIDS, autoimmune diseases, infections, atherosclerosis, ischemia-reperfusion injury, and cancer, suggesting these ectoenzymes are novel therapeutic targets for managing a variety of disorders.

Nitric oxide: a novel link between synaptic and nonsynaptic transmission

Accumulating evidence indicates that nitric oxide (NO) inhibits the function of monoamine transporters. Because the production of NO by neuronal NO synthase (nNOS) is closely related to the activation of NMDA receptors, the level of NO around nNOS-containing synapses reflects the activity of glutamate-mediated neurotransmission. Glutamate participates mainly in synaptic interactions, but with the help of NO, the strength of excitatory input might be nonsynaptically signaled to the surrounding monoaminergic neurons, which can adapt to the changes without receiving glutamatergic input and without synthesizing glutamate receptors. Thus, the effect of NO on transporters represents a new form of interneuronal communication, a nonsynaptic interaction without receptors.

Preferential release of ATP and its extracellular catabolism as a source of adenosine upon high- but not low-frequency stimulation of rat hippocampal slices

Abstract: The release of adenosine and ATP evoked by electrical field stimulation in rat hippocampal slices was investigated with the following two patterns of stimulation: (1) a brief, high‐frequency burst stimulation (trains of stimuli at 100 Hz for 50 ms applied every 2 s for 1 min), to mimic a long‐term potentiation (LTP) stimulation paradigm, and (2) a more prolonged (3 min) and low‐frequency (5 Hz) train stimulation, to mimic a long‐term depression (LTD) stimulation paradigm. The release of ATP was greater at a brief, high‐frequency burst stimulation, whereas the release of [3H]adenosine was slightly greater at a more prolonged and low‐frequency stimulation. To investigate the source of extracellular adenosine, the following two pharmacological tools were used; α,β‐methylene ADP (AOPCP), an inhibitor of ecto‐5′‐nucleotidase, to assess the contribution of the catabolism of released adenine nucleotides as a source of extracellular adenosine, and S‐(4‐nitrobenzyl)‐6‐thioinosine (NBTI), an inhibitor of adenosine transporters, to assess the contribution of the release of adenosine, as such, as a source of extracellular adenosine. At low‐frequency stimulation, NBTI inhibited by nearly 50% the evoked outflow of [3H]adenosine, whereas AOPCP inhibited [3H]adenosine outflow only marginally. In contrast, at high‐frequency stimulation, AOPCP inhibited by 30% the evoked release of [3H]adenosine, whereas NBTI produced a 40% inhibition of [3H]adenosine outflow. At both frequencies, the kinetics of evoked [3H]adenosine outflow was affected in different manners by AOPCP and NBTI; NBTI mainly depressed the rate of evoked [3H]adenosine outflow, whereas AOPCP mainly inhibited the later phase of evoked [3H]adenosine accumulation. These results show that there is a simultaneous, but quantitatively different, release of ATP and adenosine from rat hippocampal slices stimulated at frequencies that can induce plasticity phenomena such as LTP (100 Hz) or LTD (5 Hz). The source of extracellular adenosine is also different according to the frequency of stimulation; i.e., at a brief, high‐frequency stimulation there is a greater contribution of released adenine nucleotides for the formation of extracellular adenosine than at a low frequency with a more prolonged stimulation.

Involvement of Cannabinoid Receptors in the Regulation of Neurotransmitter Release in the Rodent Striatum: A Combined Immunochemical and Pharmacological Analysis

Despite the profound effect of cannabinoids on motor function, and their therapeutic potential in Parkinsons and Huntingtons diseases, the cellular and subcellular distributions of striatal CB1 receptors are not well defined. Here, we show that CB1 receptors are primarily located on GABAergic (vesicular GABA transporter-positive) and glutamatergic [vesicular glutamate transporter-1 (VGLUT-1)- and VGLUT-2-positive] striatal nerve terminals and are present in the presynaptic active zone, in the postsynaptic density, as well as in the extrasynaptic membrane. Both the nonselective agonist WIN55212-2 [(R)-(+)-[2,3-dihydro-5-methyl-3[(4-morpholinyl)methyl] pyrrolo[1,2,3-de]-1,4-benzoxazinyl]-(1-naphthalenyl)methanone mesylate salt] (EC50, 32 nm) and the CB1-selective agonist ACEA [N-(2-chloroethyl)-5Z,8Z,11Z,14Z-eicosatetraenamide] inhibited [3H]GABA release from rat striatal slices. The effect of these agonists was prevented by the CB1-selective antagonists SR141716A [N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide] (1 μm) and AM251 [1-(2,4-dichlorophenyl)-5-(4-iodophenyl)-4-methyl-N-1-piperidinyl-1H-pyrazole-3-carboxamide trifluoroacetate salt] (1 μm), indicating that cannabinoids inhibit the release of GABA via activation of presynaptic CB1 receptors. Cannabinoids modulated glutamate release via both CB1 and non-CB1 mechanisms. Cannabinoid agonists and antagonists inhibited 25 mm K+-evoked [3H]glutamate release and sodium-dependent [3H]glutamate uptake. Partial involvement of CB1 receptors is suggested because low concentrations of SR141716A partly and AM251 fully prevented the effect of WIN55212-2 and CP55940 [5-(1,1-dimethylheptyl)-2-[5-hydroxy-2-(3-hydroxypropyl)cyclohexyl]phenol]. However, the effect of CB1 agonists and antagonists persisted in CB1 knock-out mice, indicating the involvement of non-CB1,CB1-like receptors. In contrast, cannabinoids did not modulate [3H]dopamine release or [3H]dopamine and [3H]GABA uptake. Our results indicate distinct modulation of striatal GABAergic and glutamatergic transmission by cannabinoids and will facilitate the understanding of the role and importance of the cannabinoid system in normal and pathological motor function.

Involvement of P2X7 receptors in the regulation of neurotransmitter release in the rat hippocampus.

Although originally cloned from rat brain, the P2X7 receptor has only recently been localized in neurones, and functional responses mediated by these neuronal P2X7 receptors (P2X7R) are largely unknown. Here we studied the effect of P2X7R activation on the release of neurotransmitters from superfused rat hippocampal slices. ATP (1–30 mm) and other ATP analogues elicited concentration‐dependent [3H]GABA outflow, with the following rank order of potency: benzoylbenzoylATP (BzATP) > ATP > ADP. PPADS, the non‐selective P2‐receptor antagonist (3–30 µm), Brilliant blue G (1–100 nm) the P2X7‐selective antagonist and Zn2+ (0.1–30 µm) inhibited, whereas lack of Mg2+ potentiated the response by ATP. In situ hybridization revealed that P2X7R mRNA is expressed in the neurones of the cell body layers in the hippocampus. P2X7R immunoreactivity was found in excitatory synaptic terminals in CA1 and CA3 region targeting the dendrites of pyramidal cells and parvalbumin labelled structures. ATP (3–30 µm) and BzATP (0.6–6 µm) elicited concentration‐dependent [14C]glutamate efflux, and blockade of the kainate receptor‐mediated transmission by CNQX (10–100 µm) and gadolinium (100 µm), decreased ATP evoked [3H]GABA efflux. The Na+ channel blocker TTX (1 µm), low temperature (12°C), and the GABA uptake blocker nipecotic acid (1 mm) prevented ATP‐induced [3H]GABA efflux. Brilliant blue G and PPADS also reduced electrical field stimulation‐induced [3H]GABA efflux. In conclusion, P2X7Rs are localized to the excitatory terminals in the hippocampus, and their activation regulates the release of glutamate and GABA from themselves and from their target cells.

Adenosine Augments IL-10 Production by Macrophages through an A2B Receptor-Mediated Posttranscriptional Mechanism

Adenosine receptor ligands have anti-inflammatory effects and modulate immune responses by up-regulating IL-10 production by immunostimulated macrophages. The adenosine receptor family comprises G protein-coupled heptahelical transmembrane receptors classified into four types: A1, A2A, A2B, and A3. Our understanding of the signaling mechanisms leading to enhanced IL-10 production following adenosine receptor occupancy on macrophages is limited. In this study, we demonstrate that adenosine receptor occupancy increases IL-10 production by LPS-stimulated macrophages without affecting IL-10 promoter activity and IL-10 mRNA levels, indicating a posttranscriptional mechanism. Transfection experiments with reporter constructs containing sequences corresponding to the AU-rich 3′-untranslated region (UTR) of IL-10 mRNA confirmed that adenosine receptor activation acts by relieving the translational repressive effect of the IL-10 3′-UTR. By contrast, adenosine receptor activation failed to liberate the translational arrest conferred by the 3′-UTR of TNF-α mRNA. The IL-10 3′-UTR formed specific complexes with proteins present in cytoplasmic extracts of RAW 264.7 cells. Adenosine enhanced binding of proteins to a region of the IL-10 3′-UTR containing the GUAUUUAUU nonamer. The stimulatory effect of adenosine on IL-10 production was mediated through the A2B receptor, because the order of potency of selective agonists was 5′-N-ethylcarboxamidoadenosine (NECA) > N6-(3-iodobenzyl)-adenosine-5′-N-methyluronamide (IB-MECA) > 2-chloro-N6-cyclopentyladenosine (CCPA) = 2-p-(2-carboxyethyl)phenethylamino-5′-N-ethyl-carboxamidoadenosine (CGS-21680). Also, the selective A2B antagonist, alloxazine, prevented the effect of adenosine. Collectively, these studies identify a novel pathway in which activation of a G protein-coupled receptor augments translation of an anti-inflammatory gene.

Fast two-photon in vivo imaging with three-dimensional random-access scanning in large tissue volumes

The understanding of brain computations requires methods that read out neural activity on different spatial and temporal scales. Following signal propagation and integration across a neuron and recording the concerted activity of hundreds of neurons pose distinct challenges, and the design of imaging systems has been mostly focused on tackling one of the two operations. We developed a high-resolution, acousto-optic two-photon microscope with continuous three-dimensional (3D) trajectory and random-access scanning modes that reaches near-cubic-millimeter scan range and can be adapted to imaging different spatial scales. We performed 3D calcium imaging of action potential backpropagation and dendritic spike forward propagation at sub-millisecond temporal resolution in mouse brain slices. We also performed volumetric random-access scanning calcium imaging of spontaneous and visual stimulation–evoked activity in hundreds of neurons of the mouse visual cortex in vivo. These experiments demonstrate the subcellular and network-scale imaging capabilities of our system.

Modulation of lipopolysaccharide-induced tumor necrosis factor-α production by selective α- and β-adrenergic drugs in mice

In a previous study we demonstrated that mice pretreated with the highly selective alpha 2-adrenoceptor antagonist CH-38083 showed blunting of the tumor necrosis factor-alpha (TNF-alpha) response induced by bacterial lipopolysaccharide (LPS). In the present study, the effect of a selective block of alpha 2-adrenoreceptors and the role of the sympathetic nervous system (SNS) in the regulation of LPS-induced TNF-alpha production was explored further using different selective adrenoceptor antagonists and agonists. While adrenalectomy did not prevent the effect of CH-38083, the block of the sympathetic transmission by chlorisondamine fully abolished the inhibitory effect of CH-38083 on LPS-induced TNF-alpha production, suggesting that the effect of the alpha 2-adrenoceptor blocking agent is corticosteroid-independent, but it requires intact sympathetic activity. Since the selective block of alpha 2-adrenoceptors results in an increased sympathetic activity and an increase of the release of noradrenaline (NA) in both the central and the peripheral nervous systems, and in our experiments propranolol, a non-selective beta-adrenoceptor antagonist, and atenolol, a selective antagonist of beta 1-adrenoceptors, prevented the effect of alpha 2-adrenoceptor blockade by CH-38083 of the TNF-alpha response induced by LPS, it seems likely that the excessive stimulation by NA of beta 1-adrenoceptors is responsible for this action. The role of beta-adrenoceptors and endogenous catecholamines is further substantiated by the finding that pretreatment of animals with propranolol alone resulted in a dose-dependent increase of the TNF-alpha response induced by LPS, and that isoproterenol, a non-selective beta-adrenoceptor agonist, decreased it.(ABSTRACT TRUNCATED AT 250 WORDS)

A2B adenosine receptors in immunity and inflammation

A(2B) adenosine receptors are increasingly recognized as important orchestrators of inflammation. A(2B) receptor activation promotes the inflammatory response of mast cells, epithelial cells, smooth muscle cells and fibroblasts, thereby contributing to the pathophysiology of asthma and colitis. A(2B) receptor stimulation limits endothelial cell inflammatory responses and permeability and suppresses macrophage activation thereby preventing tissue injury after episodes of hypoxia and ischemia. A(2B) receptor stimulation also promotes the production of angiogenic cytokines by endothelial cells, mast cells and dendritic cells, aiding granuloma tissue formation and inflammatory resolution, but can also contribute to tumor growth. A(2B) receptors are, thus, potentially important pharmacological targets in treating immune system dysfunction and inflammation.