Miniature Excitatory Postsynaptic Currents Research Articles

Transient Enhanced GluA2 Expression in Young Hippocampal Neurons of a Fragile X Mouse Model.

AMPA-type glutamate receptors (AMPARs) are tetrameric ligand-gated channels made up of combinations of GluA1-4 subunits and play important roles in synaptic transmission and plasticity. Here, we have investigated the development of AMPAR-mediated synaptic transmission in the hippocampus of the Fmr1 knock-out (KO) mouse, a widely used model of Fragile X syndrome (FXS). FXS is the leading monogenic cause of intellectual disability and autism spectrum disorders (ASD) and it is considered a neurodevelopmental disorder. For that reason, we investigated synaptic properties and dendritic development in animals from an early stage when synapses are starting to form up to adulthood. We found that hippocampal CA1 pyramidal neurons in the Fmr1-KO mouse exhibit a higher AMPAR-NMDAR ratio early in development but reverses to normal values after P13. This increase was accompanied by a larger presence of the GluA2-subunit in synaptic AMPARs that will lead to altered Ca2+ permeability of AMPARs that could have a profound impact upon neural circuits, learning, and diseases. Following this, we found that young KO animals lack Long-term potentiation (LTP), a well-understood model of synaptic plasticity necessary for proper development of circuits, and exhibit an increased frequency of spontaneous miniature excitatory postsynaptic currents, a measure of synaptic density. Furthermore, post hoc morphological analysis of recorded neurons revealed altered dendritic branching in the KO group. Interestingly, all these anomalies are transitory and revert to normal values in older animals. Our data suggest that loss of FMRP during early development leads to temporary upregulation of the GluA2 subunit and this impacts synaptic plasticity and altering morphological dendritic branching.

Interval-dependent neurotoxicity after multiple ketamine injections in late postnatal mice.

Measuring the neurotoxic effects of multiple anesthetic exposures during neurodevelopment is complex due to the numerous factors that can affect the outcome. While we recently discovered that the interval between multiple sevoflurane exposures can affect the level of neurotoxicity, the significance of interval for other anesthetic agents is unknown. Thus, we evaluated the significance of dosing interval in the neurotoxic effects of multiple ketamine injections in postnatal day (PND) 17 mice. PND17 mice of both sexes were intraperitoneally injected with ketamine (35mg/kg) three times at short (2h) or long (24h) intervals. Changes in synaptic transmission were measured in hippocampal pyramidal neurons 5days after the last injection, and behavioral changes were assessed at the age of 8weeks. Values are presented as mean ± SD. Whereas short-interval ketamine injections enhanced excitatory synaptic transmission, as evidenced by an increased frequency of miniature excitatory postsynaptic currents (mEPSCs; ketamine, 0.09 ± 0.07Hz; control, 0.06 ± 0.03Hz), long-interval ketamine injections did not; instead, they decreased the amplitude of miniature inhibitory postsynaptic currents (mIPSCs; ketamine, 47.72 ± 6.90 pA; control, 51.21 ± 7.65 pA,). However, only long-interval ketamine injections induced long-term changes in anxiety behavioral in the open-field test (decrease in center duration; ketamine, 400.1 ± 162.8s; control, 613.3 ± 312.7s). Multiple ketamine injections induce interval-dependent, long-lasting synaptic changes and behavioral impairments. Future studies should carefully consider the dosing interval as a significant factor when studying the neurotoxic effects of multiple anesthetic exposures.

Inhibition of glutamatergic transmission and neuronal excitability by oxycodone in the rat hippocampal CA3 neurons.

Oxycodone, a semisynthetic opioid analgesic with actions similar to morphine, is extensively prescribed for treatment of moderate to severe acute pain. Given that glutamate plays a crucial role in mediating pain transmission, the purpose of this study was to investigate the effect of oxycodone on glutamatergic synaptic transmission in rat hippocampal CA3 area, which is associated with the modulation of nociceptive perception. Whole-cell patch-clamp recordings revealed that oxycodone effectively reduced presynaptic glutamate release, as detected by decreased frequencies of spontaneous excitatory postsynaptic currents (sEPSCs) and miniature EPSCs (mEPSCs), without eliciting significant changes in the amplitudes of sEPSCs and mEPSCs and glutamate-evoked inward currents. The inhibitory effect of oxycodone on the frequency of sEPSCs was blocked by the nonselective opioid receptor antagonist naloxone. In addition, oxycodone suppressed burst firing induced by 4-aminopyridine and tonic repetitive firing evoked by the applied depolarizing current. These results suggest that oxycodone inhibits spontaneous presynaptic glutamate release possibly by activating opioid receptors and consequently suppressing the neuronal excitability of hippocampal CA3 neurons.

Loss of Dysbindin Implicates Synaptic Vesicle Replenishment Dysregulation as a Potential Pathogenic Mechanism in Schizophrenia

The schizophrenia-susceptibility gene, dystrobrevin-binding protein 1 (DTNBP1), encodes the dysbindin protein and mediates neurotransmission and neurodevelopment in normal subjects. Functional studies show that DTNBP1 loss may cause deficient presynaptic vesicle transmission, which is related to multiple psychiatric disorders. However, the functional mechanism of dysbindin-mediated synaptic vesicle transmission has not been investigated systematically. In this study, we performed electrophysiological recordings in calyx of Held synapses. We found that excitatory postsynaptic current (EPSC) and miniature EPSC (mEPSC) amplitudes were unchanged in dysbindin-deficient synapses, but readily releasable pool (RRP) size and calcium dependent vesicle replenishment were affected during high-frequency stimulation. Moreover, dysbindin loss accompanied slightly decreases in Munc18-1 and snapin expression levels, which are associated with vesicle priming and synaptic homeostasis under high-frequency stimulation. Together, we inferred that dysbindin directly interacts with Munc18-1 and snapin to mediate calcium dependent RRP replenishment. Dysbindin loss may lead to RRP replenishment dysregulation during high-frequency stimulation, potentially causing cognitive impairment in schizophrenia.

Serotonin receptor 4 regulates hippocampal astrocyte morphology and function.

Astrocytes are an important component of the multipartite synapse and crucial for proper neuronal network function. Although small GTPases of the Rho family are powerful regulators of cellular morphology, the signaling modules of Rho-mediated pathways in astrocytes remain enigmatic. Here we demonstrated that the serotonin receptor 4 (5-HT4 R) is expressed in hippocampal astrocytes, both in vitro and in vivo. Through fluorescence microscopy, we established that 5-HT4 R activation triggered RhoA activity via Gα13 -mediated signaling, which boosted filamentous actin assembly, leading to morphological changes in hippocampal astrocytes. We investigated the effects of these 5-HT4 R-mediated changes in mixed cultures and in acute slices, in which 5-HT4 R was expressed exclusively in astrocytes. In both systems, 5-HT4 R-RhoA signaling changed glutamatergic synaptic transmission: It increased the frequency of miniature excitatory postsynaptic currents (mEPSCs) in mixed cultures and reduced the paired-pulse-ratio (PPR) of field excitatory postsynaptic potentials (fEPSPs) in acute slices. Overall, our present findings demonstrate that astrocytic 5-HT4 R-Gα13 -RhoA signaling is a previously unrecognized molecular pathway involved in the functional regulation of excitatory synaptic circuits.

Topiramate inhibits offensive aggression through targeting ventrolateral periaqueductal gray

Topiramate is an approved antiepileptic drug clinically used to treat epilepsy and prevent migraines. Currently, topiramate has been found to be effective in treating aggressive symptoms in neuropsychiatric patients. In preclinical studies, however, the effects and mechanisms of topiramate on offensive aggression are still largely uninvestigated. Our previous studies indicated that glutamatergic transmission in the ventrolateral periaqueductal gray (vlPAG) plays a crucial role in regulating elements of offensive aggressive behaviors. In the present work, we investigated the actions of topiramate on vlPAG glutamatergic transmission and aggressive behaviors in group-housed (GH) and socially isolated (SI) rats. The results suggested that a single injection of topiramate systemically and dose-dependently inhibited elements of offensive aggressive behaviors of both GH and SI rats in the resident-intruder test (RIT), with long-lasting effective time profiles in SI rats. Moreover, systemic single administration of topiramate reduced the frequency and amplitude of miniature excitatory postsynaptic currents (mEPSCs) in the vlPAG. Bath perfusion of topiramate directly decreased the frequency and amplitude of mEPSCs and shortened the amplitude of evoked excitatory postsynaptic currents (EPSCs) in the vlPAG. Furthermore, intra-vlPAG single microinjection of topiramate dose-dependently inhibited offensive aggressive behaviors in GH and SI rats in a time-dependent manner. Additionally, both systemic and local topiramate inhibited offensive aggressive behaviors in a (2R,6R)-hydroxynorketamine (HNK)-dependent rat model. In conclusion, the present results suggest that topiramate exerts anti-aggressive roles through its inhibitory actions on glutamatergic activities in the vlPAG. These preclinical results support topiramate as a candidate drug to treat patients with heightened offensive aggression.

Serotonergic Modulation of Spontaneous and Evoked Transmitter Release in Layer II Pyramidal Cells of Rat Somatosensory Cortex.

As axons from the raphe nuclei densely innervate the somatosensory cortex, we investigated how serotonin (5-HT) modulates transmitter release in layer II pyramidal cells of rat barrel cortex. In the presence of tetrodotoxin and gabazine, 10μM 5-HT caused a waxing and waning in the frequency of miniature excitatory postsynaptic currents (mEPSC) with no effect on amplitude. Specifically, within 15 min of recording the mEPSC frequency initially increased by 28 ± 7%, then dropped to below control (-15 ± 3%), before resurging back to 27 ± 7% larger than control. These changes were seen in 47% of pyramidal cells (responders) and were mediated by 5-HT2C receptors (5-HT2CR). Waxing resulted from phospholipase C activation, IP3 production, and Ca2+ release from presynaptic stores. Waning was prevented if PKC was blocked. In contrast, in paired recordings, the unitary EPSC amplitude was reduced by 50 ± 3% after 5-HT exposure in almost all cases with no significant effect on paired-pulse ratio and synaptic dynamics. This sustained EPSC reduction was also caused by 5-HT2R, but was mediated by presynaptic Gβγ subunits likely limiting influx through CaV2 channels. EPSC reduction, together with enhanced spontaneous noise in a restricted subset of inputs, could temporarily diminish the signal-to-noise ratio and affect the computation in the neocortical microcircuit.

Deletion of Kv10.2 Causes Abnormal Dendritic Arborization and Epilepsy Susceptibility.

The abnormal function of the voltage-gated potassium channel Kv10.2 can induce epilepsy. However, the physiological function of Kv10.2 in the central nervous system remains unclear. In this study, we found that Kv10.2 knockout (KO) increased the complexity of neurons in the CA3 subarea of hippocampus. Kv10.2 KO led to enlarged somata, elongated dendritic length, and increased the number of dendritic tips in cultured rat hippocampus neurons. Kv10.2 KO also increased Synapsin I and PSD95 protein density in cultured rat hippocampal neurons. Whole cell patch-clamp recordings of brain slices in the CA3 subarea of hippocampus revealed that Kv10.2 KO increased the amplitude of spontaneous excitatory postsynaptic currents (sEPSC) and miniature excitatory postsynaptic currents (mEPSC), depolarized the resting membrane potential and increased the action potential firing, reduced the rheobase and increased the input resistance, which results in enhanced neuronal excitability. Furthermore, we made electroencephalogram (EEG) recordings of brain activity in freely moving rats before and after inducing seizures by pentylenetetrazole (PTZ) injection. Kv10.2 KO rats dramatically increased the EEG amplitude during epilepsy. Behavioral observation after seizure induction revealed that Kv10.2 KO rats demonstrated shortened onset latency, prolonged duration, and increased seizure severity when compared with wild type rats. Therefore, this study provides a new link between Kv10.2 and neuronal morphology and higher intrinsic excitability.

Metformin Enhances Excitatory Synaptic Transmission onto Hippocampal CA1 Pyramidal Neurons.

Metformin (Met) is a first-line drug for type 2 diabetes mellitus (T2DM). Numerous studies have shown that Met exerts beneficial effects on a variety of neurological disorders, including Alzheimer’s disease (AD), Parkinson’s disease (PD) and Huntington’s disease (HD). However, it is still largely unclear how Met acts on neurons. Here, by treating acute hippocampal slices with Met (1 μM and 10 μM) and recording synaptic transmission as well as neuronal excitability of CA1 pyramidal neurons, we found that Met treatments significantly increased the frequency of miniature excitatory postsynaptic currents (mEPSCs), but not amplitude. Neither frequency nor amplitude of miniature inhibitory postsynaptic currents (mIPSCs) were changed with Met treatments. Analysis of paired-pulse ratios (PPR) demonstrates that enhanced presynaptic glutamate release from terminals innervating CA1 hippocampal pyramidal neurons, while excitability of CA1 pyramidal neurons was not altered. Our results suggest that Met preferentially increases glutamatergic rather than GABAergic transmission in hippocampal CA1, providing a new insight on how Met acts on neurons.

Pathway-specific dysregulation of striatal excitatory synapses by LRRK2 mutations.

LRRK2 is a kinase expressed in striatal spiny projection neurons (SPNs), cells which lose dopaminergic input in Parkinson's disease (PD). R1441C and G2019S are the most common pathogenic mutations of LRRK2. How these mutations alter the structure and function of individual synapses on direct and indirect pathway SPNs is unknown and may reveal pre-clinical changes in dopamine-recipient neurons that predispose toward disease. Here, R1441C and G2019S knock-in mice enabled thorough evaluation of dendritic spines and synapses on pathway-identified SPNs. Biochemical synaptic preparations and super-resolution imaging revealed increased levels and altered organization of glutamatergic AMPA receptors in LRRK2 mutants. Relatedly, decreased frequency of miniature excitatory post-synaptic currents accompanied changes in dendritic spine nano-architecture, and single-synapse currents, evaluated using two-photon glutamate uncaging. Overall, LRRK2 mutations reshaped synaptic structure and function, an effect exaggerated in R1441C dSPNs. These data open the possibility of new neuroprotective therapies aimed at SPN synapse function, prior to disease onset.

What Happened in the Hippocampal Axon in a Rat Model of Posttraumatic Stress Disorder.

Studies from postmortem and animal models have revealed altered synapse morphology and function in the brain of posttraumatic stress disorder (PTSD). And the effects of PTSD on dendrites and spines have been reported, however, the effection on axon include microtubule (MT) and synaptic vesicles of presynaptic elements remains unknown. Hippocampus is involved in abnormal memory in PTSD. In the present study, we used the single prolonged stress (SPS) model to mimic PTSD. Quantitative real-time polymerase chain reaction (RT-qPCR) and high-throughput sequencing (GSE153081) were utilized to analyze differentially expressed genes (DEGs) in the hippocampus of control and SPS rats. Immunofluorescence and western blotting were performed to examine change in axon-related proteins. Synaptic function was evaluated by measuring miniature excitatory postsynaptic currents (mEPSCs). RNA-sequencing analysis revealed 230 significantly DEGs between the control and SPS groups. Gene Ontology analysis revealed upregulation in axonemal assembly, MT formation, or movement, but downregulation in axon initial segment and synaptic vesicles fusion in the hippocampus of SPS rats. Increased expression in tau, β-tubulin MAP1B, KIF9, CCDC40, DNAH12 and decreased expression in p-tau, stathmin suggested SPS induced axon extension. Increased protein expression in VAMP, STX1A, Munc18-1 and decreased expression in synaptotagmin-1 suggested SPS induced more SNARE complex formation but decreased ability in synaptic vesicle fusion to presynaptic active zone membrane in the hippocampus of SPS rats. Further, low mEPSC frequency in SPS rats indicated dysfunction in presynaptic membrane. These results suggest that axon extension and synaptic vesicles fusion abnormality are involved in dysfunction of PTSD.

High-frequency spinal cord stimulation treatment attenuates the increase in spinal glutamate release and spinal miniature excitatory postsynaptic currents in rats with spared nerve injury-induced neuropathic pain

High-frequency spinal cord stimulation (HFSCS) at 10 kHz provides paresthesia-free treatment for chronic pain. However, the underlying mechanisms of its action have not been fully elucidated. The aim of the present study was to investigate the effect of HFSCS treatment on spinal glutamate release and uptake in spared nerve injury (SNI) rats. HFSCS was applied to the T10/T11 spinal cord 3 days after SNI. The concentration of spinal glutamate, glutamate transporter activity and miniature excitatory postsynaptic currents (mEPSCs) from neurons in lamina II were evaluated. HFSCS treatment alleviated SNI pain induced by mechanical and cold allodynia. HFSCS treatment also partially restored altered spinal glutamate uptake activity, the levels of spinal glutamate, and the frequency of mEPSCs following SNI. In conclusion, HFSCS treatment attenuated SNI-induced neuropathic pain and partially restored the altered glutamate uptake after SNI.

Antinociceptive effect of selective G protein-gated inwardly rectifying K+ channel agonist ML297 in the rat spinal cord.

The G protein-gated inwardly rectifying K+ (GIRK) channels play important signaling roles in the central and peripheral nervous systems. However, the role of GIRK channel activation in pain signaling remains unknown mainly due to the lack of potent and selective GIRK channel activators until recently. The present study was designed to determine the effects and mechanisms of ML297, a selective GIRK1/2 activator, on nociception in the spinal cord by using behavioral studies and whole-cell patch-clamp recordings from substantia gelatinosa (SG) neurons. Rats were prepared for chronic lumber catheterization and intrathecal administration of ML297. The nociceptive flexion reflex was tested using an analgesy-meter, and the influence on motor performance was assessed using an accelerating rotarod. We also investigated pre- and post-synaptic actions of ML297 in spinal cord preparations by whole-cell patch-clamp recordings. Intrathecal administration of ML297 increased the mechanical nociceptive threshold without impairing motor function. In voltage-clamp mode of patch-clamp recordings, bath application of ML297 induced outward currents in a dose-dependent manner. The ML297-induced currents demonstrated specific equilibrium potential like other families of potassium channels. At high concentration, ML297 depressed miniature excitatory postsynaptic currents (mEPSCs) but not their amplitude. The ML297-induced outward currents and suppression of mEPSCs were not inhibited by naloxone, a μ-opioid receptor antagonist. These results demonstrated that intrathecal ML297 showed the antinociceptive effect, which was mediated through direct activation of pre- and post-synaptic GIRK channels. Selective GIRK channel activation is a promising strategy for the development of new agents against chronic pain and opioid tolerance.

Physiology and Morphological Correlates of Excitatory Transmission are Preserved in Glutamine Transporter SN1-Depleted Mouse Frontal Cortex

Glutamine is an astroglia-derived precursor of the neurotransmitter glutamate, and its astroglia-to-neuron transfer is controlled by distinct glutamine transporters on the astrocytic and neuronal sites. In this study, we focused on the role of astrocytic glutamine efflux-mediating system N transporter SN1 in the maintenance of glutamatergic neurotransmission by analyzing the electrophysiological parameters ex vivo in the brain slices from control mice and mice in which vivo-morpholino technique was used to diminish SN1 protein. The glutamatergic transmission was characterized by electrophysiological recordings, ultrastructure of neuron terminals, and determination of proteins related to glutamate synaptic transmission: synaptophysin, synaptotagmin, and vit1A. The space-restricted ∼51,5% reduction of SN1 protein did not affect the expression of the neuronal glutamine transporter SAT2. SN1 depletion resulted in a reduction of field potentials (FPs), unaltered frequency of spontaneous and miniature excitatory postsynaptic currents (sEPSCs/mEPSCs), and presented a tendency towards a decrease of long-term potentiation (LTP). Ultrastructurally, preserved number of synaptic vesicles, primarily localized centrally of the cell body, correlates with unchanged levels of synaptic proteins. Collectively, the study indicates that glutamatergic transmission proceeds relatively independently of the SN1 - mediated glutamine transfer to the synapse.

The two faces of synaptic failure in AppNL-G-F knock-in mice

BackgroundIntensive basic and preclinical research into Alzheimer’s disease (AD) has yielded important new findings, but they could not yet been translated into effective therapies. One of the reasons is the lack of animal models that sufficiently reproduce the complexity of human AD and the response of human brain circuits to novel treatment approaches. As a step in overcoming these limitations, new App knock-in models have been developed that avoid transgenic APP overexpression and its associated side effects. These mice are proposed to serve as valuable models to examine Aß-related pathology in “preclinical AD.”MethodsSince AD as the most common form of dementia progresses into synaptic failure as a major cause of cognitive deficits, the detailed characterization of synaptic dysfunction in these new models is essential. Here, we addressed this by extracellular and whole-cell patch-clamp recordings in AppNL-G-F mice compared to AppNL animals which served as controls.ResultsWe found a beginning synaptic impairment (LTP deficit) at 3–4 months in the prefrontal cortex of AppNL-G-F mice that is further aggravated and extended to the hippocampus at 6–8 months. Measurements of miniature EPSCs and IPSCs point to a marked increase in excitatory and inhibitory presynaptic activity, the latter accompanied by a moderate increase in postsynaptic inhibitory function.ConclusionsOur data reveal a marked impairment of primarily postsynaptic processes at the level of synaptic plasticity but the dominance of a presumably compensatory presynaptic upregulation at the level of elementary miniature synaptic function.

Glucose Overload Inhibits Glutamatergic Synaptic Transmission: A Novel Role for CREB-Mediated Regulation of Synaptotagmins 2 and 4.

Glucose metabolism derangement is critically involved in the age-related memory loss but the underlying molecular mechanisms are still poorly understood. In a mouse model of type 1 diabetes we found memory impairment associated with inhibition of the transcription factor CREB and alteration of pre- and post-synaptic protein expression in the hippocampus. Accordingly, glucose excess negatively affected activity-dependent CREB phosphorylation and CREB-mediated mRNA expression of synaptic proteins in hippocampal primary neurons. Specifically, glucose excess inhibited the activity-dependent recruitment of CREB on the regulatory sequences of synaptotagmin (SYT) 2 and 4 promoters and the expression of SYT4 protein. As a result, high glucose affected both the frequency of miniature excitatory postsynaptic currents and NMDA receptor-mediated currents in autaptic hippocampal neuronal cultures. Collectively, our findings highlight novel mechanisms underlying hyperglycaemia-related memory loss, including CREB-dependent downregulation of synaptotagmin expression.

Cannabidiol inhibits febrile seizure by modulating AMPA receptor kinetics through its interaction with the N-terminal domain of GluA1/GluA2

Cannabidiol (CBD) is a major phytocannabinoid in Cannabis sativa. CBD is being increasingly reported as a clinical treatment for neurological diseases. Febrile seizure is one of the most common diseases in children with limited therapeutic options. We investigated possible therapeutic effects of CBD on febrile seizures and the underlying mechanism. Use of a hyperthermia-induced seizures model revealed that CBD significantly prolonged seizure latency and reduced the severity of thermally-induced seizures. Hippocampal neuronal excitability was significantly decreased by CBD. Further, CBD significantly reduced the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) mediated evoked excitatory postsynaptic currents (eEPSCs) and the amplitude and frequency of miniature EPSCs (mEPSCs). Furthermore, CBD significantly accelerated deactivation in GluA1 and GluA2 subunits. Interestingly, CBD slowed receptor recovery from desensitization of GluA1, but not GluA2. These effects on kinetics were even more prominent when AMPAR was co-expressed with γ-8, the high expression isoform 8 of transmembrane AMPAR regulated protein (TARPγ8) in the hippocampus. The inhibitory effects of CBD on AMPAR depended on its interaction with the distal N-terminal domain of GluA1/GluA2. CBD inhibited AMPAR activity and reduced hippocampal neuronal excitability, thereby improving the symptoms of febrile seizure in mice. The putative binding site of CBD in the N-terminal domain of GluA1/GluA2 may be a drug target for allosteric gating modulation of AMPAR.

Inflammation induces developmentally regulated sumatriptan inhibition of spinal synaptic transmission.

While triptans are used to treat migraine, there is evidence that they also reduce inflammation-induced pain at the spinal level. The cellular mechanisms underlying this spinal enhancement are unknown. We examined whether inflammation alters sumatriptan modulation of synaptic transmission in the rat spinal dorsal horn. Three to four days following intraplantar injection of complete Freund's adjuvant (CFA) or saline, whole cell recordings of evoked glutamatergic EPSCs were made from lumbar lamina I-II dorsal horn neurons in rat spinal slices KEY RESULTS: In 2- to 3-week-old animals, sumatriptan reduced the amplitude of evoked EPSCs and this was greater in slices from CFA, compared to saline-injected rats. In CFA-injected animals, sumatriptan increased the paired pulse ratio of evoked EPSCs and reduced the rate of spontaneous miniature EPSCs. The 5-HT1B and 5-HT1D agonists CP9 3129 and PNU109291 both inhibited evoked EPSCs in CFA but not saline-injected rats. By contrast, the 5-HT1A agonist R(+)-8-OH-DPAT inhibited evoked EPSCs in saline but not CFA-injected rats. In CFA-injected rats, the sumatriptan-induced inhibition of evoked EPSCs was reduced by the 5-HT1B and 5-HT1D antagonists NAS181 and BRL-15572. Intriguingly, the difference in sumatriptan inhibition between CFA and saline-injected animals was only observed in animals less than 4 weeks old. These findings indicate that inflammation induces a developmentally regulated 5-HT1B/1D presynaptic inhibition of excitatory transmission into the rat superficial dorsal horn. Thus, triptans could potentially act as spinal analgesic agents for inflammatory pain in the juvenile setting.

Effect of Alpha 1-Adrnoceptor Antagonists on Postsynaptic Sensitivity in Substantia Gelatinosa Neurons From Lumbosacral Spinal Cord in Rats Using Slice Patch-Clamp Technique for mEPSC.

PurposeAlpha1-adrenoceptors participate in improving storage symptoms of male lower urinary tract symptoms (LUTS). However, the mechanism of action of these compounds remains unclear. To clarify the mechanism of the α1-adrenoceptor antagonists, the amplitude of miniature excitatory postsynaptic currents (mEPSCs) was analyzed in the lumbosacral spinal cord in rats.MethodsMale adult Sprague-Dawley rats were used. Blind whole-cell patch-clamp recordings were performed on substantia gelatinosa (SG) neurons in spinal cord slice preparations. The amplitude of mEPSCs was recorded in individual SG neurons to which α1-adrenoceptors (100μM naftopidil, 100μM tamsulosin, and 30μM silodosin) were applied sequentially with intervening washout periods. Individual amplitudes were analyzed.ResultsPearson correlation coefficients (r) for the amplitudes of mEPSCs between the baseline and postadministration of α1-adrenoceptor antagonists indicated changes of the amplitude ranked in the order of naftopidil (r =0.393), tamsulosin (r=0.738), and silodosin (r=0.944). Together, the α1-adrenoceptor antagonists yielded significant increases in the amplitude of mEPSCs in SG neurons (n=108, P=0.012). However, the effects of each α1-adrenoceptor antagonist on the amplitude were as follows (relative to the baseline; n=36 each): naftopidil, P=0.129; tamsulosin, P=0.201; and silodosin, P=0.005. The rate of response to naftopidil for the outward current was relatively high among the α1-adrenoceptor blockers. An inward current was observed only with the naftopidil application.ConclusionsAlpha1-adrenoceptor antagonists changed the amplitudes of mEPSCs in a subset of SG neurons in slices prepared from the L6–S1 levels of rat spine. Although the α1-adrenoceptor antagonists generated inward or outward currents in the SG neurons, different rates of response were observed with each antagonist. These results are important for understanding the mechanisms of action (at the spinal level) of α1-adrenoceptor antagonists for the storage symptoms of male LUTS.

Cortical neurons derived from human pluripotent stem cells lacking FMRP display altered spontaneous firing patterns

BackgroundFragile X syndrome (FXS), a neurodevelopmental disorder, is a leading monogenetic cause of intellectual disability and autism spectrum disorder. Notwithstanding the extensive studies using rodent and other pre-clinical models of FXS, which have provided detailed mechanistic insights into the pathophysiology of this disorder, it is only relatively recently that human stem cell-derived neurons have been employed as a model system to further our understanding of the pathophysiological events that may underlie FXS. Our study assesses the physiological properties of human pluripotent stem cell-derived cortical neurons lacking fragile X mental retardation protein (FMRP).MethodsElectrophysiological whole-cell voltage- and current-clamp recordings were performed on two control and three FXS patient lines of human cortical neurons derived from induced pluripotent stem cells. In addition, we also describe the properties of an isogenic pair of lines in one of which FMR1 gene expression has been silenced.ResultsNeurons lacking FMRP displayed bursts of spontaneous action potential firing that were more frequent but shorter in duration compared to those recorded from neurons expressing FMRP. Inhibition of large conductance Ca2+-activated K+ currents and the persistent Na+ current in control neurons phenocopies action potential bursting observed in neurons lacking FMRP, while in neurons lacking FMRP pharmacological potentiation of voltage-dependent Na+ channels phenocopies action potential bursting observed in control neurons. Notwithstanding the changes in spontaneous action potential firing, we did not observe any differences in the intrinsic properties of neurons in any of the lines examined. Moreover, we did not detect any differences in the properties of miniature excitatory postsynaptic currents in any of the lines.ConclusionsPharmacological manipulations can alter the action potential burst profiles in both control and FMRP-null human cortical neurons, making them appear like their genetic counterpart. Our studies indicate that FMRP targets that have been found in rodent models of FXS are also potential targets in a human-based model system, and we suggest potential mechanisms by which activity is altered.