Excitatory Nerve Terminals Research Articles

Characterization of β-Hydroxybutyrate as a Cell Autonomous Fuel for Active Excitatory and Inhibitory Neurons: β-Hydroxybutyrate as a Fuel for Active Neurons.

The ketogenic diet is an effective treatment for drug-resistant epilepsy, but the therapeutic mechanisms are poorly understood. Although ketones are able to fuel the brain, it is not known whether ketones are directly metabolized by neurons on a time scale sufficiently rapid to fuel the bioenergetic demands of sustained synaptic transmission. Here, we show that nerve terminals can use the ketone β-hydroxybutyrate in a cell- autonomous fashion to support neurotransmission in both excitatory and inhibitory nerve terminals and that this flexibility relies on Ca2+ dependent upregulation of mitochondrial metabolism. Using a genetically encoded ATP sensor, we show that inhibitory axons fueled by ketones sustain much higher ATP levels under steady state conditions than excitatory axons, but that the kinetics of ATP production following activity are slower when using ketones as fuel compared to lactate/pyruvate for both excitatory and inhibitory neurons.

Phospholipid scramblase Xkr8 is required for developmental axon pruning via phosphatidylserine exposure

The mature mammalian brain connectome emerges during development via the extension and pruning of neuronal connections. Glial cells have been identified as key players in the phagocytic elimination of neuronal synapses and projections. Recently, phosphatidylserine has been identified as neuronal “eat‐me” signal that guides elimination of unnecessary input sources, but the associated transduction systems involved in such pruning are yet to be described. Here, we identified Xk‐related protein 8 (Xkr8), a phospholipid scramblase, as a key factor for the pruning of axons in the developing mammalian brain. We found that mouse Xkr8 is highly expressed immediately after birth and required for phosphatidylserine exposure in the hippocampus. Mice lacking Xkr8 showed excess excitatory nerve terminals, increased density of cortico‐cortical and cortico‐spinal projections, aberrant electrophysiological profiles of hippocampal neurons, and global brain hyperconnectivity. These data identify phospholipid scrambling by Xkr8 as a central process in the labeling and discrimination of developing neuronal projections for pruning in the mammalian brain.

Control of a hippocampal recurrent excitatory circuit by cannabinoid receptor-interacting protein Gap43

The type-1 cannabinoid receptor (CB1R) is widely expressed in excitatory and inhibitory nerve terminals, and by suppressing neurotransmitter release, its activation modulates neural circuits and brain function. While the interaction of CB1R with various intracellular proteins is thought to alter receptor signaling, the identity and role of these proteins are poorly understood. Using a high-throughput proteomic analysis complemented with an array of in vitro and in vivo approaches in the mouse brain, we report that the C-terminal, intracellular domain of CB1R interacts specifically with growth-associated protein of 43 kDa (GAP43). The CB1R-GAP43 interaction occurs selectively at mossy cell axon boutons, which establish excitatory synapses with dentate granule cells in the hippocampus. This interaction impairs CB1R-mediated suppression of mossy cell to granule cell transmission, thereby inhibiting cannabinoid-mediated anti-convulsant activity in mice. Thus, GAP43 acts as a synapse type-specific regulatory partner of CB1R that hampers CB1R-mediated effects on hippocampal circuit function.

Narrative Review of the Mechanisms and Treatment of Cough in Asthma, Cough Variant Asthma, and Non-asthmatic Eosinophilic Bronchitis.

Chronic cough is a debilitating condition affecting 10-12% of the general population and is one of the leading causes for referral to secondary care. Many conditions have been associated with chronic cough, including asthma, gastro-esophageal reflux disease and upper airways cough syndrome. Inflammatory airway conditions including cough variant asthma (CVA) and non-asthmatic eosinophilic bronchitis (NAEB) contribute to a significant proportion of presentations with chronic cough, with differing diagnostic criteria and different responses to commonly used asthma therapy for their respective diagnoses. Mechanistic studies in both animal models and humans have identified increased neuronal sensitivity and subsequent central sensitization. These mechanisms include inflammatory-mediated nociceptor sensitization and alterations of afferent nerve terminal excitability, phenotypic changes in the vagal afferent neurons over time, and central neuroplasticity resulting from increased synaptic signalling from peripheral afferents. The aim of this review is to discuss the mechanisms, neurophysiology, and management approaches currently available for patients presenting with chronic cough with underlying asthma, CVA, and NAEB and to shed a light on areas of further research required to elucidate the mechanisms of cough in this patient population.

Distinct synaptic vesicle recycling in inhibitory nerve terminals is coordinated by SV2A.

Proper brain function requires a balance between excitatory and inhibitory neuronal activity. This balance, which is disrupted in various neural disorders, ultimately depends on the functional properties of both excitatory and inhibitory neurons; however, how the physiological properties of presynaptic terminals are controlled in these neurons is largely unknown. In this study, we generated pHluorin-conjugated, synaptic vesicle-specific tracers that are preferentially expressed in excitatory or inhibitory nerve terminals. We found that synaptic vesicle recycling is ∼1.8-fold slower in inhibitory nerve terminals than excitatory nerve terminals, resulting in reduced efficacy of synaptic transmission in inhibitory presynaptic terminals during repetitive activities. Interestingly, this relative difference in trafficking efficiency is mediated by synaptic vesicle protein 2A (SV2A), which is more highly expressed in inhibitory synapses and differentially controls sorting of synaptic protein, synaptotagmin I. These findings indicate that SV2A coordinates distinct properties of synaptic vesicle recycling between excitatory and inhibitory synapses.

Effects of triphenyltin on glycinergic transmission on rat spinal neurons

Glycine is a fast inhibitory transmitter like γ-aminobutyric acid in the mammalian spinal cord and brainstem, and it is involved in motor reflex, nociception, and neuronal development. Triphenyltin (TPT) is an organometallic compound causing environmental hazard to many wild creatures. Our previous findings show that TPT ultimately induces a drain and/or exhaustion of glutamate in excitatory presynaptic nerve terminals, resulted in blockage of neurotransmission as well as methylmercury. Therefore, we have investigated the neurotoxic mechanism how TPT modulates inhibitory glycinergic transmission in the synaptic bouton preparation of rat isolated spinal neurons using a patch clamp technique. TPT at environmentally relevant concentrations (3–300 nM) significantly increased the number of frequency of glycinergic spontaneous and miniature inhibitory postsynaptic currents (sIPSC and mIPSC) without affecting the current amplitude and decay time. The TPT effects were also observed in external Ca2+-free solution containing tetrodotoxin (TTX) but removed in Ca2+-free solution with both TTX and BAPTA-AM (Ca2+ chelator). On the other hand, the amplitude of glycinergic evoked inhibitory postsynaptic currents (eIPSCs) increased with decreasing failure rate (Rf) and paired pulse ratio (PPR) in the presence of 300 nM TPT. At a high concentration (1 µM), TPT completely blocked eIPSCs after a transient facilitation. Overall, these results suggest that TPT directly acts transmitter-releasing machinery in glycinergic nerve terminals. Effects of TPT on the nerve terminals releasing fast transmitters were greater in the order of glycinergic > glutamatergic > GABAergic ones. Thus, TPT is supposed to cause a strong synaptic modulations on glycinergic neurotransmission in wild creatures.

Calcium dependence of spontaneous neurotransmitter release.

Spontaneous release of neurotransmitters is regulated by extracellular [Ca2+ ] and intracellular [Ca2+ ]. Curiously, some of the mechanisms of Ca2+ signaling at central synapses are different at excitatory and inhibitory synapses. While the stochastic activity of voltage-activated Ca2+ channels triggers a majority of spontaneous release at inhibitory synapses, this is not the case at excitatory nerve terminals. Ca2+ release from intracellular stores regulates spontaneous release at excitatory and inhibitory terminals, as do agonists of the Ca2+ -sensing receptor. Molecular machinery triggering spontaneous vesicle fusion may differ from that underlying evoked release and may be one of the sources of heterogeneity in release mechanisms.

Spike and Neuropeptide-Dependent Mechanisms Control GnRH Neuron Nerve Terminal Ca2+ over Diverse Time Scales.

Fast cell-to-cell communication in the brain is achieved by action potential-dependent synaptic release of neurotransmitters. The fast kinetics of transmitter release are determined by transient Ca2+ elevations in presynaptic nerve terminals. Neuromodulators have previously been shown to regulate transmitter release by inhibiting presynaptic Ca2+ influx. Few studies to date have demonstrated the opposite, that is, neuromodulators directly driving presynaptic Ca2+ rises and increases in nerve terminal excitability. Here we use GCaMP Ca2+ imaging in brain slices from mice to address how nerve terminal Ca2+ is controlled in gonadotropin-releasing hormone (GnRH) neurons via action potentials and neuromodulators. Single spikes and bursts of action potentials evoked fast, voltage-gated Ca2+ channel-dependent Ca2+ elevations. In contrast, brief exposure to the neuropeptide kisspeptin-evoked long-lasting Ca2+ plateaus that persisted for tens of minutes. Neuropeptide-mediated Ca2+ elevations were independent of action potentials, requiring Ca2+ entry via voltage-gated Ca2+ channels and transient receptor potential channels in addition to release from intracellular store mechanisms. Together, these data reveal that neuromodulators can exert powerful and long-lasting regulation of nerve terminal Ca2+ independently from actions at the soma. Thus, GnRH nerve terminal function is controlled over disparate timescales via both classical spike-dependent and nonclassical neuropeptide-dependent mechanisms.SIGNIFICANCE STATEMENT Nerve terminals are highly specialized regions of a neuron where neurotransmitters and neurohormones are released. Many neuroendocrine neurons release neurohormones in long-duration bursts of secretion. To understand how this is achieved, we have performed live Ca2+ imaging in the nerve terminals of gonadotropin-releasing hormone neurons. We find that bursts of action potentials and local neuropeptide signals are both capable of evoking large increases in nerve terminal Ca2+ Increases in Ca2+ driven by spike bursts last seconds; however, the increases in nerve terminal Ca2+ driven by neuropeptides can persist for tens of minutes. These findings reveal new mechanisms by which neuroendocrine nerve terminal Ca2+ can be controlled in the brain.

Putative transmembrane transporter modulates higher-level aggression in Drosophila

By selection of winners of dyadic fights for 35 generations, we have generated a hyperaggressive Bully line of flies that almost always win fights against the parental wild-type Canton-S stock. Maintenance of the Bully phenotype is temperature dependent during development, with the phenotype lost when flies are reared at 19 °C. No similar effect is seen with the parent line. This difference allowed us to carry out RNA-seq experiments and identify a limited number of genes that are differentially expressed by twofold or greater in the Bullies; one of these was a putative transmembrane transporter, CG13646, which showed consistent and reproducible twofold down-regulation in Bullies. We examined the causal effect of this gene on the phenotype with a mutant line for CG13646, and with an RNAi approach. In all cases, reduction in expression of CG13646 by approximately half led to a hyperaggressive phenotype partially resembling that seen in the Bully flies. This gene is a member of a very interesting family of solute carrier proteins (SLCs), some of which have been suggested as being involved in glutamine/glutamate and GABA cycles of metabolism in excitatory and inhibitory nerve terminals in mammalian systems.

Retrograde fibroblast growth factor 22 (FGF22) signaling regulates insulin-like growth factor 2 (IGF2) expression for activity-dependent synapse stabilization in the mammalian brain.

Communication between pre- and postsynaptic cells promotes the initial organization of synaptic specializations, but subsequent synaptic stabilization requires transcriptional regulation. Here we show that fibroblast growth factor 22 (FGF22), a target-derived presynaptic organizer in the mouse hippocampus, induces the expression of insulin-like growth factor 2 (IGF2) for the stabilization of presynaptic terminals. FGF22 is released from CA3 pyramidal neurons and organizes the differentiation of excitatory nerve terminals formed onto them. Local application of FGF22 on the axons of dentate granule cells (DGCs), which are presynaptic to CA3 pyramidal neurons, induces IGF2 in the DGCs. IGF2, in turn, localizes to DGC presynaptic terminals and stabilizes them in an activity-dependent manner. IGF2 application rescues presynaptic defects of Fgf22(-/-) cultures. IGF2 is dispensable for the initial presynaptic differentiation, but is required for the following presynaptic stabilization both in vitro and in vivo. These results reveal a novel feedback signal that is critical for the activity-dependent stabilization of presynaptic terminals in the mammalian hippocampus.

Effects of Propofol on Glutamate-Induced Calcium Mobilization in Presynaptic Boutons of Rat Hippocampal Neurons

Recent reports have suggested that various general anesthetics affect presynaptic processes in the central nervous system. However, characterizations of the influence of intravenous anesthetics on neurotransmitter release from presynaptic nerve terminals (boutons) are insufficient. Because the presynaptic calcium concentration ([Ca2+]pre) regulates neurotransmitter release, we investigate the effects of the intravenous anesthetic propofol on neurotransmitter release by measuring [Ca2+]pre in the presynaptic boutons of individual dissociated hippocampal neurons. Brain slices were prepared from Sprague–Dawley rats (10 - 14 days of age). The hippocampal CA1 area was isolated with a fire-polished glass pipette, which vibrated horizontally to dissociate hippocampal CA1 neurons along with their attached presynaptic boutons. Presynaptic boutons were visualized under a confocal laser scanning microscope after staining with FM1-43 dye, and [Ca2+]pre was measured using fluo-3 AM dye. Glutamate (3 – 100 μM) administration increased [Ca2+]pre in Ca2+- containing external solution in a concentration-dependent manner. Propofol (3 – 30 μM) dose-dependently suppressed this glutamate (30 μM)-induced increase in [Ca2+]pre in boutons attached to dendrites, but not to the soma or base of the dendritic tree. The large majority of excitatory synapses on CA1 neurons are located on dendritic spines; therefore, propofol may affect glutamate-induced Ca2+ mobilization in excitatory, but not inhibitory, presynaptic boutons. Propofol may possibly have some effect on glutamate-regulated neurotransmitter release from excitatory presynaptic nerve terminals through inhibiting the increase in [Ca2+]pre induced by glutamate.

Regulatory Effects of 5-Hydroxytryptamine Receptors on Voiding Function.

A growing body of evidence suggests that 5-hydroxytryptamine (5-HT; serotonin) has both physiological and pathological functions in the lower urinary tract. A wide variety of 5-HT receptor subtypes are variably expressed in different organs, both peripheral and central. On urinary bladder smooth muscle, 5-HT1A, 5-HT2, 5-HT3, and 5-HT7 subtypes could function as postjunctional receptors. Postjunctional 5-HT2 receptors induce detrusor contraction of the bladder body. 5-HT1A is suggested to have a similar effect to 5-HT2, while 5-HT3 might suppress detrusor contraction evoked by direct muscle stimulation. Postjunctional 5-HT7 is reported to induce relaxation of the bladder neck, which might be required for efficient voiding. 5-HT1A, 5-HT2A, 5-HT2C, 5-HT3, 5-HT4, and 5-HT7 subtypes also could act as prejunctional receptors in autonomic excitatory nerve terminals. 5-HT2A, 5-HT2C, 5-HT3, 5-HT4, and 5-HT7 subtypes facilitate the neurogenic contraction of the detrusor by enhancing cholinergic or purinergic transmission, whereas 5-HT1A receptors might inhibit the release of acetylcholine in the detrusor. Furthermore, 5-HT1D could be involved in the suppression of ATP release from the urothelium, aiding visceral sensation of the urinary bladder. In the central pathways controlling the micturition reflex, 5-HT1A, 5-HT2A, and 5-HT7 are involved in regulation of bladder and urethral sphincter activities. Their functions, especially that of 5-HT1A, vary in a species- and site (spinal or supraspinal)- dependent manner. In addition to urinary bladder, 5-HT could be involved in prostate contraction and cell proliferation. Evidence indicates that 5-HT receptor subtypes may be novel therapeutic targets for lower urinary tract symptoms. Grants-in-Aid for Scientific Research (C) (KAKENHI 23590707, 24590722, and 26460694) from the Japan Society for the Promotion of Science.

Inhibitory properties underlying non-monotonic input-output relationship in low-frequency spherical bushy neurons of the gerbil.

Spherical bushy cells (SBCs) of the anteroventral cochlear nucleus (AVCN) receive input from large excitatory auditory nerve (AN) terminals, the endbulbs of Held, and mixed glycinergic/GABAergic inhibitory inputs. The latter have sufficient potency to block action potential firing in vivo and in slice recordings. However, it is not clear how well the data from slice recordings match the inhibition in the intact brain and how it contributes to complex phenomena such as non-monotonic rate-level functions (RLF). Therefore, we determined the input-output relationship of a model SBC with simulated endbulb inputs and a dynamic inhibitory conductance constrained by recordings in brain slice preparations of hearing gerbils. Event arrival times from in vivo single-unit recordings in gerbils, where 70% of SBC showed non-monotonic RLF, were used as input for the model. Model output RLFs systematically changed from monotonic to non-monotonic shape with increasing strength of tonic inhibition. A limited range of inhibitory synaptic properties consistent with the slice data generated a good match between the model and recorded RLF. Moreover, tonic inhibition elevated the action potentials (AP) threshold and improved the temporal precision of output functions in a SBC model with phase-dependent input conductance. We conclude that activity-dependent, summating inhibition contributes to high temporal precision of SBC spiking by filtering out weak and poorly timed EPSP. Moreover, inhibitory parameters determined in slice recordings provide a good estimate of inhibitory mechanisms apparently active in vivo.

Shelter from the Glutamate storm: Loss of olivocochlear efferents increases cochlear nerve degeneration during aging

The olivocochlear (OC) feedback pathways include one population, the medial (M)OC projections to outer hair cells, which forms a sound-evoked inhibitory reflex that can reduce sound-induced cochlear vibrations, and a second population, the lateral (L)OC projections to the synaptic zone underneath the inner hair cells, that can modulate the excitability of the cochlear nerve terminals. Although there is ample evidence of OC-mediated protective effects from both of these systems when the ear is exposed to intense noise, the functional significance of this protection is questionable in a pre-industrial environment where intense noise was not so commonplace. We have re-evaluated the phenomenon of OC-mediated protection in light of recent work showing that acoustic exposure destroys cochlear neurons at sound pressure levels previously considered atraumatic, because they cause no permanent hair cell loss or threshold shift. We have shown that loss of OC innervation at a young age causes the cochlea to age a greatly accelerated rate, even without purposeful noise exposure, when aging is measured by the loss of synaptic connections between cochlear nerve fibers and hair cells. Possible relevance to hearing-in-noise problems of the elderly will be discussed.

Tanshinone IIA, a constituent of Danshen, inhibits the release of glutamate in rat cerebrocortical nerve terminals

Ethnopharmacological relevanceDanshen is a commonly used traditional Chinese medicine and has received considerable attention due to their beneficial effects on the health, including prevention of cardiovascular disease, and cancer. Tanshinone IIA, a major active constituent of Danshen, has been reported to have a neuroprotective profile. Aim of the studyAn excessive release of glutamate is considered to be related to neuropathology of several neurological diseases. In this study, we investigated whether tanshinone IIA could affect endogenous glutamate release and explored the possible mechanism. Materials and methodsThe experimental model was the isolated nerve terminals (synaptosomes) purified from the rat cerebral cortex. The release of glutamate was evoked by the K+ channel blocker 4-aminopyridine (4-AP) and measured by one-line enzyme-coupled fluorometric assay. We also used a membrane potential-sensitive dye to assay nerve terminal excitability and depolarization, and a Ca2+ indicator, Fura-2-acetoxymethyl ester, to monitor cytosolic Ca2+ concentrations ([Ca2+]C). ResultsTanshinone IIA inhibited the release of glutamate evoked by 4-AP in a concentration-dependent manner. Inhibition of glutamate release by tanshinone IIA was prevented by the chelating the extracellular Ca2+ ions, and by the vesicular transporter inhibitor bafilomycin A1. However, the glutamate transporter inhibitor DL-threo-beta-benzyl-oxyaspartate did not have any effect on the action of tanshinone IIA. Tanshinone IIA decreased the depolarization-induced increase in [Ca2+]C, whereas it did not alter the resting synaptosomal membrane potential or 4-AP-mediated depolarization. Furthermore, the effect of tanshinone IIA on evoked glutamate release was prevented by the Cav2.2 (N-type) and Cav2.1 (P/Q-type) channel blocker ω-conotoxin MVIIC, but not by the ryanodine receptor blocker dantrolene or the mitochondrial Na+/Ca2+ exchanger blocker CGP37157. Mitogen-activated protein kinase (MEK) inhibition also prevented the inhibitory effect of tanshinone IIA on evoked glutamate release. ConclusionThese results show that tanshinone IIA inhibits glutamate release from cortical synaptosomes in rats through the suppression of presynaptic voltage-dependent Ca2+ entry and MEK signaling cascade.

Presynaptic Glycine Receptors Increase GABAergic Neurotransmission in Rat Periaqueductal Gray Neurons

The periaqueductal gray (PAG) is involved in the central regulation of nociceptive transmission by affecting the descending inhibitory pathway. In the present study, we have addressed the functional role of presynaptic glycine receptors in spontaneous glutamatergic transmission. Spontaneous EPSCs (sEPSCs) were recorded in mechanically dissociated rat PAG neurons using a conventional whole-cell patch recording technique under voltage-clamp conditions. The application of glycine (100 µM) significantly increased the frequency of sEPSCs, without affecting the amplitude of sEPSCs. The glycine-induced increase in sEPSC frequency was blocked by 1 µM strychnine, a specific glycine receptor antagonist. The results suggest that glycine acts on presynaptic glycine receptors to increase the probability of glutamate release from excitatory nerve terminals. The glycine-induced increase in sEPSC frequency completely disappeared either in the presence of tetrodotoxin or Cd2+, voltage-gated Na+, or Ca2+ channel blockers, suggesting that the activation of presynaptic glycine receptors might depolarize excitatory nerve terminals. The present results suggest that presynaptic glycine receptors can regulate the excitability of PAG neurons by enhancing glutamatergic transmission and therefore play an important role in the regulation of various physiological functions mediated by the PAG.

Comparative effects of pentobarbital on spontaneous and evoked transmitter release from inhibitory and excitatory nerve terminals in rat CA3 neurons

Pentobarbital (PB) modulates GABA(A) receptor-mediated postsynaptic responses through various mechanisms, and can directly activate the channel at higher doses. These channels exist both pre- and postsynaptically, and on the soma outside the synapse. PB also inhibits voltage-dependent Na⁺ and Ca²⁺ channels to decrease excitatory synaptic transmission. Just how these different sites of action combine to contribute to the overall effects of PB on inhibitory and excitatory synaptic transmission is less clear. To compare these pre- and postsynaptic actions of PB, we used a 'synaptic bouton' preparation of isolated rat hippocampal CA3 pyramidal neurons where we could measure in single neurons the effects of PB on spontaneous and single bouton evoked GABAergic inhibitory and glutamatergic excitatory postsynaptic currents (sIPSCs, sEPSCs, eIPSCs and eEPSCs), respectively. Low (sedative) concentrations (3-10 μM) of PB increased the frequency and amplitude of sIPSCs and sEPSCs, and also presynaptically increased the amplitude of both eIPSCs and eEPSCs. There was no change in current kinetics at this low concentration. At higher concentrations (30-300 μM), PB decreased the frequency, and increased the amplitude of sIPSCs, and presynaptically decreased the amplitude of eIPSCs. The current decay phase of sIPSCs and eIPSCs was increased. An increase in both frequency and amplitude was seen for sEPSCs, while the eIPSCs was also decreased by a bicuculline-sensitive presynaptic effect. The results confirm the multiple sites of action of PB on inhibitory and excitatory transmission and demonstrate that the most sensitive site of action is on transmitter release, via effects on presynaptic GABA(A) receptors. At low concentrations, however, both glutamate and GABA release is similarly enhanced, making the final effects on neuronal excitability difficult to predict and dependent on the particular systems involved and/or on subtle differences in susceptibility amongst individuals. At higher concentrations, release of both transmitters is decreased, while the postsynaptic effects to increase IPSPs and decrease EPSCs would be expected to both results in reduced neuronal excitability.

Valproate causes reduction of the excitatory amino acid aspartate in nerve terminals

Valproate is well established in the treatment of epilepsy and psychiatric disorders, yet the main mechanism of action remains to be determined. Here we show that valproate may reduce neurotransmission of the excitatory amino acid, aspartate. By electron microscopic immunogold cytochemistry we demonstrate a 63–68% reduction in the level of aspartate in excitatory nerve terminals at 30min after an acute dose of valproate. The level of glutamate in the same terminals was unchanged by valproate treatment. In inhibitory terminals, valproate caused a 65% decrease in the aspartate level, whereas the GABA level was not significantly changed. In summary, the present study shows that valproate reduces the nerve terminal content of the excitatory neurotransmitter aspartate. This points to a new mechanism of action for valproate: reduced neuronal excitation through reduced aspartergic neurotransmission.

Neurotoxin A2NTX Blocks Fast Inhibitory and Excitatory Transmitter Release From Presynaptic Terminals

Our recent study showed a possibility that newly developed A2 type botulinum toxin (A2NTX) inhibits both spontaneous and evoked transmitter release from inhibitory (glycinergic or GABAergic) and excitatory (glutamatergic) nerve terminals using rat spinal sacral dorsal commissural nucleus neurons. In the present study, to determine the modulatory effect of A2NTX on glycinergic and glutamatergic release probabilities, we tested the effects of A2NTX on a single inhibitory or excitatory nerve ending adherent to a dissociated neuron that was activated by paired-pulse stimuli by using the focal electrical stimulation technique. The results of the present paired-pulse experiments showed clearly that A2NTX enhanced paired-pulse facilitation of evoked glycinergic inhibitory postsynaptic currents and glutamatergic excitatory postsynaptic currents and increased the failure rate (Rf) of the first postsynaptic currents (P1) and both the responses. These effects of A2NTX on the amplitude and Rf of the P1 and the second postsynaptic currents (P2) and paired-pulse ratio were rescued by application of 4-aminophthalimide. In summary, the present results showed that A2NTX acts purely presynaptically and inhibits the release machinery of transmitters such as glycine and glutamate, and the transmitter release machinery became less sensitive to intracellular free-Ca2+ in A2NTX poisoned nerve terminals.

Fibroblast Growth Factor 22 Contributes to the Development of Retinal Nerve Terminals in the Dorsal Lateral Geniculate Nucleus

At least three forms of signaling between pre- and postsynaptic partners are necessary during synapse formation. First, “targeting” signals instruct presynaptic axons to recognize and adhere to the correct portion of a postsynaptic target cell. Second, trans-synaptic “organizing” signals induce differentiation in their synaptic partner so that each side of the synapse is specialized for synaptic transmission. Finally, in many regions of the nervous system an excess of synapses are initially formed, therefore “refinement” signals must either stabilize or destabilize the synapse to reinforce or eliminate connections, respectively. Because of both their importance in processing visual information and their accessibility, retinogeniculate synapses have served as a model for studying synaptic development. Molecular signals that drive retinogeniculate “targeting” and “refinement” have been identified, however, little is known about what “organizing” cues are necessary for the differentiation of retinal axons into presynaptic terminals. To identify such “organizing” cues, we used microarray analysis to assess whether any target-derived “synaptic organizers” were enriched in the mouse dorsal lateral geniculate nucleus (dLGN) during retinogeniculate synapse formation. One candidate “organizing” molecule enriched in perinatal dLGN was FGF22, a secreted cue that induces the formation of excitatory nerve terminals in muscle, hippocampus, and cerebellum. In FGF22 knockout mice, the development of retinal terminals in dLGN was impaired. Thus, FGF22 is an important “organizing” cue for the timely development of retinogeniculate synapses.