N-Acetylaspartylglutamate: the most abundant peptide neurotransmitter in the mammalian central nervous system

In the progress of science, as in life, timing is important. The acidic dipeptide, N-acetylaspartylglutamate (NAAG), was discovered in the mammalian nervous system in 1965, but initially was not considered to be a neurotransmitter candidate. In the mid-1980s, a few laboratories revisited the question of NAAG's role in the nervous system and pursued hypotheses regarding its function that ranged from a precursor for the transmitter pool of glutamate to a direct role as a peptide transmitter. Since that time, NAAG has been tested against nearly all of the established criteria for identification of a neurotransmitter. It successfully meets each of these tests, including a concentrated presence in neurons and synaptic vesicles, release from axon endings in a calcium-dependent manner following initiation of action potentials, and extracellular hydrolysis by membrane-bound peptidase activity. NAAG is the most prevalent and widely distributed neuropeptide in the mammalian nervous system. NAAG activates NMDA receptors with a low potency that may vary among receptor subtypes, and it is a highly selective agonist at the type 3 metabotropic glutamate receptor (mGluR3). Acting through this receptor, NAAG reduces cyclic AMP levels, decreases voltage-dependent calcium conductance, suppresses excitotoxicity, influences long-term potentiation and depression, regulates GABA(A) receptor subunit expression, and inhibits synaptic release of GABA from cortical neurons. Cloning of peptidase activities against NAAG provides opportunities to study the cellular and molecular mechanisms by which synaptic NAAG peptidase activity is controlled. Given the codistribution of this peptide with a spectrum of traditional transmitters and its ability to activate mGluR3, we speculate that one role for NAAG following synaptic release is the activation of metabotropic autoreceptors that inhibit subsequent transmitter release. A second role is the production of extracellular glutamate following NAAG hydrolysis.     

Sprouting and formation of new synapses in motor structures of the central nervous system

The investigations of sprouting and reactive synaptogenesis in motor structures of the spinal cord, brain stem, thalamus, and cerebral cortex are reviewed. The reactions of the neurons and neuronal connections to injury and the ability of the nervous system to recover the impaired connections in the early postnatal period are compared with those in adult animals. The sprouting phenomenon appearing in the intact central nervous system is analyzed too. The mechanisms of synaptic reorganization of the nervous centers are discussed.Neirofiziologiya/Neurophysiology, Vol. 26, No. 4, pp. 299–314, June–July, 1994.     

Cholinergic transmission via central synapses in the locust nervous system

In this study we examine the nature of chemical synaptic transmission between identified filiform hair receptors on the prothoracic segment of a locust and the identified postsynaptic projection interneuron (A4I1). The effects of pressure ejected acetylcholine, and various ligands of acetylcholine receptors on the activity of the postsynaptic neuron A4I1, or on wind-elicited responses in A4I1 are reported. It is suggested that the transmitter of the afferent fibers is acetylcholine, and that fast transmission is mediated by nicotinic acetylcholine-receptors. Both nicotine and carbachol act as agonists, whereas d-tubocurarine and alpha-bungarotoxin act as antagonists. The presence of muscarinic acetylcholine receptors was also evident from the modulatory effects of muscarine, oxotremorine and pilocarpine, which were blocked by bath application of atropine. GABA, and its agonists muscimol and cis-4-amino-crotonic-acid lead to inhibition of A4I1 responses. This inhibition was prevented by the additional application of picrotoxin. This suggests involvement of a ligand-gated GABA receptor which, most likely, increases chloride conductance. Metabotropic GABA-receptors do not seem to be involved, since baclofene, diazepam and bicuculline ejections had no effects. Glutamate also inhibits wind elicited A4I1 responses. Although attempts were made to further characterize the receptor involved, tested substances such as kainic acid, glycine, CNQX or GDEE had no effect.     

Agrin plays an organizing role in the formation of sympathetic synapses

Agrin is a nerve-derived factor that directs neuromuscular synapse formation, however its role in regulating interneuronal synaptogenesis is less clear. Here, we examine agrin's role in synapse formation between cholinergic preganglionic axons and sympathetic neurons in the superior cervical ganglion (SCG) using agrin-deficient mice. In dissociated cultures of SCG neurons, we found a significant decrease in the number of synapses with aggregates of presynaptic synaptophysin and postsynaptic neuronal acetylcholine receptor among agrin-deficient neurons as compared to wild-type neurons. Moreover, the levels of pre- and postsynaptic markers at the residual synapses in agrin-deficient SCG cultures were also reduced, and these defects were rescued by adding recombinant neural agrin to the cultures. Similarly, we observed a decreased matching of pre- and postsynaptic markers in SCG of agrin-deficient embryos, reflecting a decrease in the number of differentiated synapses in vivo. Finally, in electrophysiological experiments, we found that paired-pulse depression was more pronounced and posttetanic potentiation was significantly greater in agrin-deficient ganglia, indicating that synaptic transmission is also defective. Together, these findings indicate that neural agrin plays an organizing role in the formation and/or differentiation of interneuronal, cholinergic synapses.     

Actions of amantadine at synaptic and extrasynaptic cholinergic receptors in the central nervous system of the cockroach Periplaneta americana

The effects of amantadine were investigated on cercal afferent, giant interneurone synapses and on the cell body membrane of the fast coxal depressor motoneurone (D_f), in the cockroach Periplaneta americana. Bath-applied amantadine at concentrations above 2.0 × 10^?5 M significantly reduced the amplitude of unitary and compound epsps recorded by sucrose-gap methods from cercal afferent, giant interneurone synapses in the desheathed sixth abdominal ganglion. Complete block of synaptic transmission was achieved at 1.0 × 10^?3 M amantadine. Synaptic blockade, which was not accompanied by changes in resting potential, was almost fully reversed by washing the ganglion in normal saline. From the dose-dependence of the synaptic blocking action, a Hill coefficient of 0.94 was estimated, indicating that there is no co-operativity in the binding of amantadine to its site of action.Bath-application of amantadine (5.0 × 10^?5 M) resulted in a parallel shift to the right of the dose-response curve for the depolarizing postsynaptic actions of acetylcholine. Nevertheless, even at a concentration of 2.0 × 10^?3 M, amantadine failed to protect the synaptic acetylcholine receptor/ion channel complex from the blocking action of α-bungarotoxin (5.0 × 10^?7 M). In addition, the block by amantadine of the acetylcholine-induced current recorded from the cell body membrane of the fast coxal depressor motoneurone (D_f), was strongly dependent on membrane potential in the range ? 120mV to ? 70mV. An action of amantadine at the open acetylcholine receptor/ion channel complex is proposed.     

Ultrastructural peculiarities of interneuronal and neuromuscular relations in the trunkal part of the intraepidermal nervous system of Nerilla Sp. (Archiannelida)

This study deals with ultrastructural analysis of interneuronal and neuromuscular relations in a representative of archiannelid Nerilla sp. with primitive intraepidermal type of the nervous system. A particular attention has been paid to the area of ventral ciliate groove and the associated site of epidermis. In the ciliate groove, sensory and motor cilia are revealed and described. Sites of axonal terminals of the sensory cells supplied with cilia are noted in the epidermal nerve plexus. Epidermal-muscular cells and nerve terminals on them are revealed. Various interneuronal contact variants both of non-synaptic and of typically synaptic types are described. An attention is drawn to the rare presence of contacts of the gap junction type among interneuronal contacts in Nerilla sp. In sufficiently differentiated synapses of the chemical type, phenomena of exocytosis are described. There are shown specific features of innervation of longitudinal (somatic) musculature of the neril-lid body, including input of synaptic vesicles into the basal lamina substance and their translocation into the depth of the muscular layer.     

Real-time measurements of vesicle-SNARE recycling in synapses of the central nervous system

Following the fusion of synaptic vesicles with the presynaptic plasma membrane of nerve terminals by the process of exocytosis, synaptic-vesicle components are recycled to replenish the vesicle pool. Here we use a pH-sensitive green fluorescent protein to measure the residence time of VAMP, a vesicle-associated SNARE protein important for membrane fusion, on the surfaces of synaptic terminals of hippocampal neurons following exocytosis. The time course of VAMP retrieval depends linearly on the amount of VAMP that is added to the plasma membrane, with retrieval occurring between about 4 seconds and 90 seconds after exocytosis, and newly internalized vesicles are rapidly acidified. These data are well described by a model in which endocytosis appears to be saturable, but proceeds with an initial maximum velocity of about one vesicle per second. We also find that, following exocytosis, a portion of the newly inserted VAMP appears on the surface of the axon.     

Ultrastructure of synapses in the central nervous system of lamellibranch molluscs.

Fine-structural characteristics of synaptic contacts were investigated in the central nervous system of different species of lamellibranch molluscs. Neuropile of the ganglia is characterized by regular occurrence of ultrastructurally well-defined polarized chemical synapses resembling those described in other invertebrate species and vertebrates. In addition to the generally observed membrane thickenings, enhanced density of synaptic membranes, cleft material and vesicle clustering on the presynaptic membrane, synapses are occasionally characterized by other and pinocytotic invaginations. Synaptic connections were distinguished on the basis of the vesicle content of the presynaptic terminal. Different forms of synaptic configurations (divergence, convergence, presynaptic modification) were observed in the ganglia.     

Amine sulfate formation in the central nervous system

The sulfates of norepinephrine, dopamine (DA), and serotonin (5-hydroxytryptamine [5HT]) are present in the cerebrospinal fluid (CSF) of laboratory animals and humans. The amounts of sulfated amines in human CSF always greatly exceed the amounts of the free amines. The enzyme responsible for sulfation, phenol sulfotransferase (PST) (EC 2.8.2.1), has been detected in the brain tissue of several species, including humans. PST in the human brain has a high affinity for the amines but it is a low-capacity enzyme. Accordingly, sulfation appears to be of greater significance in the economy of the amines under quiescent conditions than during conditions of increased release of transmitter. Recent evidence suggests that a fraction of the conjugated amines in CSF enters from plasma because in the African green monkey, DA sulfate and 5HT sulfate cross the blood-CSF barrier after i.v. injection. In addition, in humans there are no increases in the concentration of amine sulfates from lumbar to ventricular CSF that would also be compatible with a partly peripheral origin for the amine sulfates. However, it appears that at least a portion of the amine sulfates in CSF originate in the central nervous system because the ratios of [CSF amine sulfates]/[plasma sulfates] are never as high after i.v. injection as under basal conditions.     

Comparison of opioid receptor distributions in the rat central nervous system

The opioid receptors, mu, delta and kappa, conduct the major pharmacological effects of opioid drugs, and exhibit intriguing functional relationships and interactions in the CNS. Previously established hypotheses regarding the mechanisms underlying these phenomena specify theoretical patterns of relative cellular localisation for the different receptor types. In this study, we have used double-label immunohistochemistry to compare the cellular distributions of delta and kappa receptors with those of mu receptors in the rat CNS. Regions of established significance in opioid addiction were examined. Extensive mu/delta co-localisation was observed in neuron-like cells in several regions. mu and kappa receptors were also often co-localised in neuron-like cell bodies in several regions. However, intense kappa immunoreactivity (ir) also appeared in a separate, morphologically distinct population of cells that did not express mu receptors. These small, ovoid cells were often closely apposed against the larger, mu-ir cell bodies. Such cellular appositions were seen in several regions, but were particularly common in the medial thalamus, the periaqueductal grey and brainstem regions. These findings support proposals that functional similarities, synergy and cooperativity between mu and delta receptors arise from widespread co-expression by cells and intracellular molecular interactions. Although co-expression of mu and kappa receptors was also detected, the appearance of a separate population of kappa-expressing cells supports proposals that the contrasting and functionally antagonistic properties of mu and kappa receptors are due to expression in physiologically distinct cell types. Greater understanding of opioid receptor interaction mechanisms may provide possibilities for therapeutic intervention in opioid addiction and other conditions.     

Fine-tuning the central nervous system: microglial modelling of cells and synapses

Microglia constitute as much as 10–15% of all cells in the mammalian central nervous system (CNS) and are the only glial cells that do not arise from the neuroectoderm. As the principal CNS immune cells, microglial cells represent the first line of defence in response to exogenous threats. Past studies have largely been dedicated to defining the complex immune functions of microglial cells. However, our understanding of the roles of microglia has expanded radically over the past years. It is now clear that microglia are critically involved in shaping neural circuits in both the developing and adult CNS, and in modulating synaptic transmission in the adult brain. Intriguingly, microglial cells appear to use the same sets of tools, including cytokine and chemokine release as well as phagocytosis, whether modulating neural function or mediating the brain''s innate immune responses. This review will discuss recent developments that have broadened our views of neuro-glial signalling to include the contribution of microglial cells.     

A mechanism for the observed recovery from ineffectiveness of synapses in the central nervous system

Three recent experiments have shown that synaptic connections in the central nervous system that are structurally present but functionally ineffective can be made to recover their effectiveness for exciting neurons. The common features of these experiments are briefly reviewed. A mechanism is proposed requiring three postulates that are each consistent with orthodox physiology. There is an opportunity for a quantitative treatment to test against future experimental results.     

Molecular biology of cadherins in the nervous system

Cadherins are cell-cell adhesion molecules belonging to the Ca²⁺-dependent cadherin superfamily. In the last few years the number of cadherins identified in the nervous system has increased considerably. Cadherins are integral membrane glycoproteins. They are structurally closely related and interspecies homologies are high. The function is mediated through a homophilic binding mechanism, and intracellular proteins, directly or indirectly connected to the cadherins and the cytoskeleton, are necessary for cadherin activity. Cadherins have been implicated in segregation and aggregation of tissues at early developmental stages and in growth and guidance of axons during nervous system development. These functions are modified by changes in type(s) and amount of cadherins expressed at different developmental stages. The regulatory elements guiding cadherin expression are currently being elucidated.     

Lysophospholipids and their receptors in the central nervous system

Lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P), two of the best-studied lysophospholipids, are known to influence diverse biological events, including organismal development as well as function and pathogenesis within multiple organ systems. These functional roles are due to a family of at least 11 G protein-coupled receptors (GPCRs), named LPA_1–6 and S1P_1–5, which are widely distributed throughout the body and that activate multiple effector pathways initiated by a range of heterotrimeric G proteins including G_i/o, G_12/13, G_q and G_s, with actual activation dependent on receptor subtypes. In the central nervous system (CNS), a major locus for these signaling pathways, LPA and S1P have been shown to influence myriad responses in neurons and glial cell types through their cognate receptors. These receptor-mediated activities can contribute to disease pathogenesis and have therapeutic relevance to human CNS disorders as demonstrated for multiple sclerosis (MS) and possibly others that include congenital hydrocephalus, ischemic stroke, neurotrauma, neuropsychiatric disorders, developmental disorders, seizures, hearing loss, and Sandhoff disease, based upon the experimental literature. In particular, FTY720 (fingolimod, Gilenya, Novartis Pharma, AG) that becomes an analog of S1P upon phosphorylation, was approved by the FDA in 2010 as a first oral treatment for MS, validating this class of receptors as medicinal targets. This review will provide an overview and update on the biological functions of LPA and S1P signaling in the CNS, with a focus on results from studies using genetic null mutants for LPA and S1P receptors. This article is part of a Special Issue entitled Advances in Lysophospholipid Research.     

The role of agrin in synaptic development,plasticity and signaling in the central nervous system

Development of the neuromuscular junction (NMJ) requires secretion of specific isoforms of the proteoglycan agrin by motor neurons. Secreted agrin is widely expressed in the basal lamina of various tissues, whereas a transmembrane form is highly expressed in the brain. Expression in the brain is greatest during the period of synaptogenesis, but remains high in regions of the adult brain that show extensive synaptic plasticity. The well-established role of agrin in NMJ development and its presence in the brain elicited investigations of its possible role in synaptogenesis in the brain. Initial studies on the embryonic brain and neuronal cultures of agrin-null mice did not reveal any defects in synaptogenesis. However, subsequent studies in culture demonstrated inhibition of synaptogenesis by agrin antisense oligonucleotides or agrin siRNA. More recently, a substantial loss of excitatory synapses was found in the brains of transgenic adult mice that lacked agrin expression everywhere but in motor neurons. The mechanisms by which agrin influences synapse formation, maintenance and plasticity may include enhancement of excitatory synaptic signaling, activation of the “muscle-specific” receptor tyrosine kinase (MuSK) and positive regulation of dendritic filopodia. In this article I will review the evidence that agrin regulates synapse development, plasticity and signaling in the brain and discuss the evidence for the proposed mechanisms.     

Differential expression of endophilin 1 and 2 dimers at central nervous system synapses

Endophilin 1 is proposed to participate in synaptic vesicle biogenesis through SH3 domain-mediated interactions with the polyphosphoinositide phosphatase synaptojanin and the GTPase dynamin. Endophilin family members have also been identified as binding partners for a number of diverse cellular proteins. We define here the endophilin 1-binding site within synaptojanin 1 and show that this sequence independently and selectively purifies from brain extracts endophilin 1 and a closely related protein, endophilin 2. Endophilin 2, like endophilin 1, is highly expressed in brain, concentrated in nerve terminals, and found in complexes with synaptojanin and dynamin. Although a fraction of endophilins 1 and 2 coexist in the same complex, the distribution of these endophilin isoforms among central synapses only partially overlaps. Endophilins 1 and 2 are found predominantly as stable dimers through a predicted coiled-coil domain in their conserved NH2-terminal moiety. Dimerization may allow endophilins to link a number of different cellular targets to the endocytic machinery.     

Development of the nervous system in hatchlings of Spadella cephaloptera (Chaetognatha), and implications for nervous system evolution in Bilateria

Chaetognaths (arrow worms) play an important role as predators in planktonic food webs. Their phylogenetic position is unresolved, and among the numerous hypotheses, affinities to both protostomes and deuterostomes have been suggested. Many aspects of their life history, including ontogenesis, are poorly understood and, though some aspects of their embryonic and postembryonic development have been described, knowledge of early neural development is still limited. This study sets out to provide new insights into neurogenesis of newly hatched Spadella cephaloptera and their development during the following days, with attention to the two main nervous centers, the brain and the ventral nerve center. These were examined with immunohistological methods and confocal laser-scan microscopic analysis, using antibodies against tubulin, FMRFamide, and synapsin to trace the emergence of neuropils and the establishment of specific peptidergic subsystems. At hatching, the neuronal architecture of the ventral nerve center is already well established, whereas the brain and the associated vestibular ganglia are still rudimentary. The development of the brain proceeds rapidly over the next 6 days to a state that resembles the adult pattern. These data are discussed in relation to the larval life style and behaviors such as feeding. In addition, we compare the larval chaetognath nervous system and that of other bilaterian taxa in order to extract information with phylogenetic value. We conclude that larval neurogenesis in chaetognaths does not suggest an especially close relationship to either deuterostomes or protostomes, but instead displays many apomorphic features.