Frontiers in Synaptic Neuroscience最新文献

Cannabis exposure during gestation evokes significant molecular modifications to neurodevelopmental programs leading to neurophysiological and behavioral abnormalities in humans. The main neuronal receptor for Δ⁹-tetrahydrocannabinol (THC) is the type-1 cannabinoid receptor CB₁R, one of the most abundant G-protein-coupled receptors in the nervous system. While THC is the major psychoactive phytocannabinoid, endocannabinoids (eCBs) are the endogenous ligands of CB₁R and are known to act as retrograde messengers to modulate synaptic plasticity at different time scales in the adult brain. Accumulating evidence indicates that eCB signaling through activation of CB₁R plays a central role in neural development. During development, most CB₁R localized to axons of projection neurons, and in mice eCB signaling impacts axon fasciculation. Understanding of eCB-mediated structural plasticity during development, however, requires the identification of the precise spatial and temporal dynamics of CB₁R-mediated modifications at the level of individual neurons in the intact brain. Here, the cell-autonomous role of CB₁R and the effects of CB₁R-mediated eCB signaling were investigated using targeted single-cell knockdown and pharmacologic treatments in Xenopus. We imaged axonal arbors of retinal ganglion cells (RGCs) in real time following downregulation of CB₁R via morpholino (MO) knockdown. We also analyzed RGC axons with altered eCB signaling following treatment with URB597, a selective inhibitor of the enzyme that degrades Anandamide (AEA), or JZL184, an inhibitor of the enzyme that blocks 2-Arachidonoylglycerol (2-AG) hydrolysis, at two distinct stages of retinotectal development. Our results demonstrate that CB₁R knockdown impacts RGC axon branching at their target and that differential 2-AG and AEA-mediated eCB signaling contributes to presynaptic structural connectivity at the time that axons terminate and when retinotectal synaptic connections are made. Altering CB₁R levels through CB₁R MO knockdown similarly impacted dendritic morphology of tectal neurons, thus supporting both pre- and postsynaptic cell-autonomous roles for CB₁R-mediated eCB signaling.

The unitary postsynaptic response to presynaptic quantal glutamate release is the fundamental basis of excitatory information transfer between neurons. The view, however, of individual glutamatergic synaptic connections in a population as homogenous, fixed-strength units of neural communication is becoming increasingly scrutinized. Here, we used minimal stimulation of individual glutamatergic afferent axons to evoke single synapse resolution postsynaptic responses from central sensory lamina I neurons in an ex vivo adult rat spinal slice preparation. We detected unitary events exhibiting a NMDA receptor component with distinct kinetic properties across synapses conferred by specific GluN2 subunit composition, indicative of GluN2 subtype-based postsynaptic heterogeneity. GluN2A, 2A and 2B, or 2B and 2D synaptic predominance functioned on distinct lamina I neuron types to narrowly, intermediately, or widely tune, respectively, the duration of evoked unitary depolarization events from resting membrane potential, which enabled individual synapses to grade differentially depolarizing steps during temporally patterned afferent input. Our results lead to a model wherein a core locus of proteomic complexity prevails at this central glutamatergic sensory synapse that involves distinct GluN2 subtype configurations. These findings have major implications for subthreshold integrative capacity and transmission strength in spinal lamina I and other CNS regions.

The efficiency of neocortical information processing critically depends on the balance between the glutamatergic (excitatory, E) and GABAergic (inhibitory, I) synaptic transmission. A transient imbalance of the E/I-ratio during early development might lead to neuropsychiatric disorders later in life. The transgenic glutamic acid decarboxylase 67-green fluorescent protein (GAD67-GFP) mouse line (KI) was developed to selectively visualize GABAergic interneurons in the CNS. However, haplodeficiency of the GAD67 enzyme, the main GABA synthetizing enzyme in the brain, temporarily leads to a low GABA level in the developing brain of these animals. However, KI mice did not demonstrate any epileptic activity and only few and mild behavioral deficits. In the present study we investigated how the developing somatosensory cortex of KI-mice compensates the reduced GABA level to prevent brain hyperexcitability. Whole-cell patch clamp recordings from layer 2/3 pyramidal neurons at P14 and at P21 revealed a reduced frequency of miniature inhibitory postsynaptic currents (mIPSCs) in KI mice without any change in amplitude or kinetics. Interestingly, mEPSC frequencies were also decreased, while the E/I-ratio was nevertheless shifted toward excitation. Surprisingly, multi-electrode-recordings (MEA) from acute slices revealed a decreased spontaneous neuronal network activity in KI mice compared to wild-type (WT) littermates, pointing to a compensatory mechanism that prevents hyperexcitability. Blockade of GABA_B receptors (GABA_BRs) with CGP55845 strongly increased the frequency of mEPSCs in KI, but failed to affect mIPSCs in any genotype or age. It also induced a membrane depolarization in P14 KI, but not in P21 KI or WT mice. MEA recordings in presence of CGP55845 revealed comparable levels of network activity in both genotypes, indicating that tonically activated GABA_BRs balance neuronal activity in P14 KI cortex despite the reduced GABA levels. Blockade of GABA transporter 3 (GAT-3) reproduced the CGP55845 effects suggesting that tonic activation of GABA_BRs is mediated by ambient GABA released via GAT-3 operating in reverse mode. We conclude that GAT-3-mediated GABA release leads to tonic activation of both pre- and postsynaptic GABA_BRs and restricts neuronal excitability in the developing cortex to compensate for reduced neuronal GABA synthesis. Since GAT-3 is predominantly located in astrocytes, GAD67 haplodeficiency may potentially stimulate astrocytic GABA synthesis through GAD67-independent pathways.

Introduction: Cancer patients treated with paclitaxel often develop chemotherapy-induced peripheral neuropathy, which has not been effectively treated with drugs. The anti-diabetic drug metformin is effective in the treatment of neuropathic pain. The aim of this study was to elucidate effect of metformin on paclitaxel-induced neuropathic pain and spinal synaptic transmission.

Methods: Electrophysiological experiments on rat spinal slices were performed in vitro and mechanical allodynia quantified in vitro.

Results: The present data demonstrated that intraperitoneal injection of paclitaxel produced mechanical allodynia and potentiated spinal synaptic transmission. Intrathecal injection of metformin significantly reversed the established mechanical allodynia induced by paclitaxel in rats. Either spinal or systemic administration of metformin significantly inhibited the increased frequency of spontaneous excitatory postsynaptic currents (sEPSCs) in spinal dorsal horn neurons from paclitaxel-treated rats. We found that 1 h incubation of metformin also reduced the frequency rather than the amplitude of sEPSCs in the spinal slices from paclitaxel-treated rats.

Discussion: These results suggested that metformin was able to depress the potentiated spinal synaptic transmission, which may contribute to alleviating the paclitaxel-induced neuropathic pain.

Since the discovery of neurexins (Nrxns) as essential and evolutionarily conserved synaptic adhesion molecules, focus has largely centered on their functional contributions to glutamatergic synapses. Recently, significant advances to our understanding of neurexin function at GABAergic synapses have revealed that neurexins can play pleiotropic roles in regulating inhibitory synapse maintenance and function in a brain-region and synapse-specific manner. GABAergic neurons are incredibly diverse, exhibiting distinct synaptic properties, sites of innervation, neuromodulation, and plasticity. Different classes of GABAergic neurons often express distinct repertoires of Nrxn isoforms that exhibit differential alternative exon usage. Further, Nrxn ligands can be differentially expressed and can display synapse-specific localization patterns, which may contribute to the formation of a complex trans-synaptic molecular code that establishes the properties of inhibitory synapse function and properties of local circuitry. In this review, we will discuss how Nrxns and their ligands sculpt synaptic inhibition in a brain-region, cell-type and synapse-specific manner.

Four modes of endocytosis and subsequent synaptic vesicle (SV) recycling have been described at the presynapse to ensure the availability of SVs for synaptic release. However, it is unclear to what extend these modes operate under physiological activity patterns in vivo. The coat protein clathrin can regenerate SVs either directly from the plasma membrane (PM) via clathrin-mediated endocytosis (CME), or indirectly from synaptic endosomes by SV budding. Here, we examined the role of clathrin in SV recycling under physiological conditions by applying the clathrin inhibitor Pitstop-2 to the calyx of Held, a synapse optimized for high frequency synaptic transmission in the auditory brainstem, in vivo. The effects of clathrin-inhibition on SV recycling were investigated by serial sectioning scanning electron microscopy (S³EM) and 3D reconstructions of endocytic structures labeled by the endocytosis marker horseradish peroxidase (HRP). We observed large endosomal compartments as well as HRP-filled, black SVs (bSVs) that have been recently recycled. The application of Pitstop-2 led to reduced bSV but not large endosome density, increased volumes of large endosomes and shifts in the localization of both types of endocytic compartments within the synapse. These changes after perturbation of clathrin function suggest that clathrin plays a role in SV recycling from both, the PM and large endosomes, under physiological activity patterns, in vivo.

Membrane lipid rafts are sphingolipids and cholesterol-enriched membrane microdomains, which form a center for the interaction or assembly of palmitoylated signaling molecules, including Src family non-receptor type protein tyrosine kinases. Lipid rafts abundantly exist in neurons and function in the maintenance of synapses. Excitatory synaptic strength is largely controlled by the surface expression of α-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) receptors in the mammalian brain. AMPA receptor endocytosis from the synaptic surface is regulated by phosphorylation of the GluA2 subunit at tyrosine 876 by Src family kinases. Here, I revealed that tyrosine phosphorylated GluA2 is concentrated in the lipid rafts fraction. Furthermore, stimulation-induced upregulation of GluA2 tyrosine phosphorylation is disrupted by the treatment of neurons with a cholesterol-depleting compound, filipin III. These results indicate the importance of lipid rafts as enzymatic reactive sites for AMPA receptor tyrosine phosphorylation and subsequent AMPA receptor internalization from the synaptic surface.