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Objective:To investigate the effect of ginkgo biloba extract (GBE) on oxidative stress in medial prefrontal cortex and excitatory/inhibitory balance of pyramidal neurons in chronic unpredictable mild stress (CUMS)-induced depressive model mice.Methods:Totally 48 SPF grade 7-week-old male C57BL/6J mice were divided into 4 groups according to random number table method: control+ saline group (CTRL+ Veh), control+ GBE group (CTRL+ GBE), model+ saline group (CUMS+ Veh), model+ GBE group (CUMS+ GBE), with 12 mice in each group.Mice in CUMS+ Veh group and CUMS+ GBE group were established by CUMS method, and mice in CTRL+ GBE group and CUMS+ GBE group were intraperitoneally injected with GBE (70 mg/kg) once a day, and mice in CTRL+ Veh group and CUMS+ Veh group were injected intraperitoneally with 0.9% sodium chloride solution.Then, the sucrose preference test, forced swimming test (FST) and tail suspension test (TST) were performed to evaluate the depressive-like behavior of mice, and open field test (OFT) was performed to evaluate the autonomous locomotion and exploration ability and anxiety-like behavior.The content of malondialdehyde (MDA) and activity of superoxide dismutase (SOD) in mPFC were determined by ELISA.Spontaneous excitatory postsynaptic currents (sEPSC) and spontaneous inhibitory postsynaptic currents (sIPSC) were detected by whole-cell recording.SPSS 23.0 was used for data analysis and two-factor analysis of variance(whether to get GBE, whether to mold, show as GBE×CUMS) was used for statistical analysis.Results:(1) Behavioral results: the the time spent in center and total distance of OFT and sugar preference rate of the four groups of mice were compared, and the interaction of GBE×CUMS was significant( F=24.90, 4.82, 3.91, all P<0.05). The results of simple effect analysis showed that the time spent in center ((47.15±3.58) s), the total distance((19.33±0.86) m) and the sugar preference rate((59.11±8.79)%) of the mice in CUMS+ Veh group were lower than those in the CTRL+ Veh group((61.55±2.49) s, (23.24±1.21) m, (84.02±7.45) %) (all P<0.01), and the above indexes in CUMS+ GBE group ((56.51±3.53) s, (20.75±1.31) m, (70.80±11.79)%) were higher than those in CUMS+ Veh group (all P<0.05). In the immobility time of FST and TST of mice in the 4 groups, the interaction of GBE×CUMS were significant( F=85.53, 83.39, both P<0.01). The immobility time of FST and TST in CUMS+ Veh group were higher than those in CTRL+ Veh group (both P<0.01 ), and the above indexes in CUMS+ GBE group were lower than CUMS+ Veh group(both P<0.05). (2)The results of ELISA showed that the interaction of GBE×CUMS of SOD level of mice in the 4 groups was not significant ( F=3.52, P=0.07), but the main effects of GBE factor and CUMS factor were both significant ( F=4.69, 46.93, both P<0.05). The interaction of GBE×CUMS of MDA level was significant( F=16.61, P<0.01). The level of SOD in the CUMS+ Veh group was lower than that in the CTRL+ Veh group ( P<0.01), and the level of SOD in the CUMS+ GBE group was higher than that in the CUMS+ Veh group ( P<0.05). The level of MDA in the CUMS+ Veh group was higher than that of the CTRL+ Veh group ( P<0.01), and the level of MDA in CUMS+ GBE group was lower than that of the CUMS+ Veh group ( P<0.01). (3) The results of whole-cell recording showed that the interaction of GBE×CUMS of frequency and quantification of sEPSC in the four groups were significant ( F=5.45, 6.94, both P<0.05). The sEPSC frequency and quantification in the CUMS+ Veh group were lower than those in the CTRL+ Veh group (both P<0.01), and the sEPSC frequency and quantification in CUMS+ GBE group were higher than those of CUMS+ Veh group (both P<0.05). The interaction of GBE×CUMS of frequency and quantification of sIPSC in the four groups were significant ( F=7.78, 8.96, both P<0.01). The sIPSC frequency and quantification of the CUMS+ Veh group were higher than those of CTRL+ Veh group (both P<0.01), and the above indexes of CUMS+ GBE group were lower than those of CUMS+ Veh group (both P<0.01). As for the sEPSC/sIPSC ratio, GBE×CUMS interaction was significant ( F=5.45, P=0.02). The sEPSC/sIPSC ratio of CUMS+ Veh group (0.09±0.01) was lower than that of CTRL+ Veh group (0.28±0.04) ( P<0.01), and the sEPSC/sIPSC ratio of CUMS+ GBE group (0.14±0.03) was higher than that of CUMS+ Veh group ( P<0.05). Conclusion:Ginkgo biloba extract can improve the depression-like behavior of mice induced by CUMS, reduce the oxidative stress of mPFC and improve the excitation/inhibition balance of pyramidal neurons in depressive model mice.
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PURPOSE: The aim of this study was to characterize the responsiveness of miniature excitatory postsynaptic currents (mEPSCs) to α1-adrenoceptor blockers in substantia gelatinosa (SG) neurons from the spinal cord to develop an explanation for the efficacy of α1-adrenoceptor blockers in micturition dysfunction. METHODS: Male adult Sprague-Dawley rats were used. Blind whole-cell patch-clamp recordings were performed using SG neurons in spinal cord slices. Naftopidil (100μM), tamsulosin (100μM), or silodosin (30μM), α1-adrenoceptor blockers, was perfused. The frequency of mEPSCs was recorded in an SG neuron to which the 3 blockers were applied sequentially with wash-out periods. Individual frequencies in a pair before naftopidil and tamsulosin perfusion were plotted as baseline, and the correlation between them was confirmed by Spearman correlation coefficient; linear regression was then performed. The same procedure was performed before naftopidil and silodosin perfusion. Frequencies of pairs after naftopidil and tamsulosin perfusion and after naftopidil and silodosin perfusion were similarly analyzed. The ratios of the frequencies after treatment to before were then calculated. RESULTS: After the treatments, Spearman ρ and the slope were decreased to 0.682 from 0.899 at baseline and 0.469 from 1.004 at baseline, respectively, in the tamsulosin group relative to the naftopidil group. In the silodosin group, Spearman ρ and the slope were also decreased to 0.659 from 0.889 at baseline and 0.305 from 0.989 at baseline, respectively, relative to the naftopidil group. Naftopidil significantly increased the ratio of the frequency of mEPSCs compared to tamsulosin and silodosin (P=0.015 and P=0.004, respectively). CONCLUSIONS: There was a difference in responsiveness in the frequency of mEPSCs to α1-adrenoceptor blockers, with the response to naftopidil being the greatest among the α1-adrenoceptor blockers. These data are helpful to understand the action mechanisms of α1-adrenoceptor blockers for male lower urinary tract symptoms in clinical usage.
Subject(s)
Adult , Animals , Humans , Male , Rats , Adrenergic alpha-1 Receptor Antagonists , Excitatory Postsynaptic Potentials , Linear Models , Lower Urinary Tract Symptoms , Neurons , Perfusion , Rats, Sprague-Dawley , Spinal Cord , Substantia Gelatinosa , UrinationABSTRACT
Objective • To investigate the mechanism of spinal chemokine C-C motif receptor 2 (CCR2)-mediated maintenance of bone cancer pain (BCP) in rats. Methods • Fifty-four SD rats were divided into BCP group, sham operation group, BCP+INCB3344 (CCR2 specific antagonist) group, and BCP+vehicle control group. Walker256 breast cancer cells were injected into the tibia medullary cavity of rats in the BCP group to establish the BCP model, while the rats in the sham operation group were injected with the same amount of saline. The rats in the BCP+INCB3344 group received intrathecal injection of INCB3344 on the 14th day after the establishment of BCP model, while the BCP+vehicle control group rats were injected with the same amount of vehicle. The mechanical pain thresholds of BCP group rats and sham operation group rats were measured to judge the success of BCP model. The expressions of CCR2 in the dorsal horn of spinal cord in the sham operation group rats and the BCP group rats were detected by Western blotting. The effects of intrathecal administration of INCB3344 on the mechanical pain threshold of BCP rats were observed by mechanical pain behavior test. Whole-cell patch-clamp recordings were used to investigate the differences of spontaneous excitatory postsynaptic currents (sEPSCs), α-amino-3-hydroxy-5-methyl-4-isoxazole-propionicacid (AMPA) and N-methyl-D-aspartic acid (NMDA)-induced currents of spinal substantia gelatinosa (SG) neurons of rats in the BCP group, the BCP+INCB3344 group and the BCP+vehicle control group. Results • Compared with the sham operation group, the mechanical pain threshold of BCP group rats reduced significantly on the 14th day after operation (P=0.000), and the expression of CCR2 in ipsilateral spinal cord of BCP group rats increased significantly (P=0.009). After intrathecal injection of INCB3344 for 4 h, the mechanical pain threshold of BCP+INCB3344 group rats was significantly increased (P=0.002). The frequency and amplitude of sEPSCs and the amplitude of AMPA and NMDA-induced currents in SG neurons of BCP group rats were significantly higher than those of the sham operation group rats (all P=0.000), while intrathecal administration of INCB3344 could significantly inhibit the above-mentioned indices in the BCP+INCB3344 group (all P<0.05). In addition, extracellular perfusion of INCB3344 could also significantly inhibit the frequency (P=0.001) and amplitude (P=0.020) of sEPSCs in SG neurons in BCP rats. Conclusion • CCR2 expressing in the spinal cord mediates the enhancement of excitatory synaptic transmission efficacy in the spinal dorsal horn of BCP rats by enhancing the functions of AMPA and NMDA receptors, which may be an important mechanism for the maintenance of BCP.
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<p><b>OBJECTIVE</b>To investigate the changes in the membrane properties and synaptic stability of the rat retinal ganglion cells (RGCs) during postnatal development.</p><p><b>METHODS</b>Whole-cell patch-clamp technique was used to record the action potentials (AP) and miniature excitatory postsynaptic currents (mEPSC) of SD rat RGCs at postnatal days 7, 14 and 40. The active and passive membrane properties and the synaptic stability (measured by the amplitude, frequency, rise time and decay time of mEPSC) of the RGCs were analyzed using Patchmaster software.</p><p><b>RESULTS</b>Comparison of the RGCs in SD rats across different postnatal ages revealed significant changes in the electrophysiological characteristics of the RGCs during postnatal development. The discharge rate was significantly greater while the AP half-peak width was significantly smaller at postnatal day 15 (P15) than at P7 ( < 0.01), but were both similar between P15 and P40 (=0.086); in terms of the passive membrane properties, the membrane time constant gradually decreased during the development. The frequency of mEPSCs increased significantly over time during postnatal development ( < 0.01), but was similar between P15 and P40 rats.</p><p><b>CONCLUSIONS</b>In SD rats, the membrane properties and synaptic stability of the RGCs undergo alterations following a specific pattern, which highlights a critical period where distinct changes occur in the electrophysiological characteristics of RGCs, followed by gradual stabilization over time. Such changes in the electrophysiological characteristics represent the basic characteristics of RGCs for visual signal processing, and understanding of this mechanism may provide insights into the exact role of the RGC in visual information processing.</p>
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PURPOSE: Naftopidil ((±)-1-[4-(2-methoxyphenyl) piperazinyl]-3-(1-naphthyloxy) propan-2-ol) is prescribed in several Asian countries for lower urinary tract symptoms suggestive of benign prostatic hyperplasia. Previous animal experiments showed that intrathecal injection of naftopidil abolished rhythmic bladder contraction in vivo. Naftopidil facilitated spontaneous inhibitory postsynaptic currents in substantia gelatinosa (SG) neurons in spinal cord slices. These results suggest that naftopidil may suppress the micturition reflex at the spinal cord level. However, the effect of naftopidil on evoked excitatory postsynaptic currents (EPSCs) in SG neurons remains to be elucidated. METHODS: Male Sprague-Dawley rats at 6 to 8 weeks old were used. Whole-cell patch-clamp recordings were made using SG neurons in spinal cord slices isolated from adult rats. Evoked EPSCs were analyzed in Aδ or C fibers. Naftopidil or prazosin, an α1-adrenoceptor blocker, was perfused at 100 μM or 10 μM, respectively. RESULTS: Bath-applied 100 μM naftopidil significantly decreased the peak amplitudes of Aδ and C fiber-evoked EPSCs to 72.0%±7.1% (n=15) and 70.0%±5.5% (n=20), respectively, in a reversible and reproducible manner. Bath application of 10μM prazosin did not inhibit Aδ or C fiber-evoked EPSCs. CONCLUSIONS: The present study suggests that a high concentration of naftopidil reduces the amplitude of evoked EPSCs via a mechanism that apparently does not involve α1-adrenoceptors. Inhibition of evoked EPSCs may also contribute to suppression of the micturition reflex, together with nociceptive stimulation.
Subject(s)
Adult , Animals , Humans , Male , Rats , Animal Experimentation , Asian People , Baths , Excitatory Postsynaptic Potentials , In Vitro Techniques , Inhibitory Postsynaptic Potentials , Injections, Spinal , Lower Urinary Tract Symptoms , Nerve Fibers, Unmyelinated , Neurons , Prazosin , Prostatic Hyperplasia , Rats, Sprague-Dawley , Reflex , Spinal Cord , Substantia Gelatinosa , Urinary Bladder , UrinationABSTRACT
BACKGROUND: The medial vestibular nucleus (MVN) is involved in the reflex control of the head and eyes, and the recovery of vestibular function after vestibular injuries. This study was performed to investigate the actions of the orphan opioid (nociceptin) on the membrane conductances and synaptic transmission in rat MVN neurons. METHODS: Whole cell patch clamp recordings were carried out in the brainstem slice of neonatal rats. RESULTS: Nociceptin (2 micro M) inhibited the spontaneous discharge in the majority (83%) of MVN neurons. This inhibition was insensitive to the non-specific opioid receptor antagonist naloxone (10 micro M), but was effectively antagonized by the selective opioid receptor-like 1 (ORL1) receptor antagonist, [Nphe1] nociceptin(1-13)NH2 (3 micro M). Nociceptin had no effect on the rate or amplitude of miniature inhibitory postsynaptic currents (mIPSCs). Nociceptin induced an outward current, and which was blocked by [Nphe1] nociceptin(1-13)NH2 in MVN neurons. Outward current reversed at -81 +/- 2 mV, which was close to the K+ equilibrium potential as calculated by the Nernst equation in 6 mM extracellular potassium solution. This indicates that the action of nociceptin involves postsynaptic receptors on the MVN neurons. CONCLUSIONS: These results suggest that nociceptin modulate neuronal excitability by activating a K+ conductance in postsynaptic neurons, not by modulation of synaptic transmission in MVN neurons.
Subject(s)
Animals , Child , Humans , Rats , Brain Stem , Child, Orphaned , Head , Inhibitory Postsynaptic Potentials , Membranes , Naloxone , Neurons , Potassium , Receptors, Opioid , Reflex , Synaptic Transmission , Vestibular NucleiABSTRACT
BACKGROUND: The medial vestibular nucleus (MVN) is involved in the reflex control of the head and eyes, and the recovery of vestibular function after vestibular lesions. This study was performed to investigate the characteristics of inhibitory postsynaptic currents (IPSC) and the roles of metabotropic glutamate receptors on inhibitory synaptic transmissions. METHODS: Whole cell patch clamp recordings were carried out from MVN neurons in brainstem slice of neonatal rats. RESULTS: The frequency and amplitude of the IPSC were significantly reduced by GABAA a n t a g o n i s t bicuculline (20 microM) but were not affected by the glycine antagonist strychnine (1 microM). The baseline frequency, amplitude and decay time constant of spontaneous IPSC (sIPSC) were 4.9+/-1.8 Hz, 25.9+/-3.1 pA, 8.7+/-0.5 ms, respectively. Glutamate (1 mM) increased the frequency of sIPSC, but decreased that of the miniature IPSC (mIPSC) in MVN neurons. Such dual effects of glutamate were mimicked by group I, II metabotropic glutamate receptor (mGluR) agonist ACPD (20 microM). The specific mGluR 2, 3 agonist DCG-IV (3 microM) reduced mIPSC frequency, but did not increase sIPSC frequency. The mGluR 1, 5 agonist DHPG (100 microM) increased sIPSC and mIPSC frequency. CONCLUSIONS: These data suggest that the IPSC recorded from MVN neurons are mediated mainly by GABAA receptors and glutamate-induced modulations of inhibitory synaptic transmissions can influence the excitability of MVN neurons.
Subject(s)
Animals , Rats , Bicuculline , Brain Stem , Glutamic Acid , Glycine , Head , Inhibitory Postsynaptic Potentials , Neurons , Receptors, Metabotropic Glutamate , Reflex , Strychnine , Synaptic Transmission , Vestibular NucleiABSTRACT
GABA (-aminobutyric acid) is one of the important neurotransmitters in the central nervous system of mammals and its action is variable according to the maturation phases of neurons. The neurons of early cultural days (less than 7 days) have been used for a developing neuronal model, while the neurons of later days (over 3 weeks) used for a mature model. This study was performed to investigate the electrophysiological property of GABAergic synapses in the hippocampal neurons cultured for 10 to 14 days which are considered to be transitional period between the developing and the mature phases. Membrane potential was depolarized and a inward transmembrane current was induced by 20 M GABA infusion. Frequency and amplitude of spontaneous postsynaptic currents (PSCs) were inhibited during the GABA infusion, but decay time constant was not affected significantly. In most hippocampal neurons, no GABAergic PSCs were observed during the administration of 0.5 M TTX, 50 M APV and 10 M CNQX. In the neurons counting 25% approximately, however, small persisted PSCs showed the existence of GABAergic synapses which were blocked by 10 M bicuculine. As the functional property of isolated GABAergic synapses, amplitude of GABAergic PSCs were diminished, and decay time constants and rising times were prolonged during the 20 M GABA infusion in all recorded neurons. In conclusion, approximately 25% of the hippocmpal neurons cultured 10 to 14 days used GABA as well as glutamate as a neurotransmitter. It seems that the GABAergic synapses composed of functionally homogenous GABAA receptors act as inhibitory modulator of the excitatory signal transmission.
Subject(s)
Animals , Rats , 6-Cyano-7-nitroquinoxaline-2,3-dione , Central Nervous System , gamma-Aminobutyric Acid , Glutamic Acid , Mammals , Membrane Potentials , Neurons , Neurotransmitter Agents , Synapses , Synaptic PotentialsABSTRACT
Objective To investigate the effects of Panax notoginseng saponins(PNS)on both the excitatory and inhibitory synaptic transmission in the pyramidal neurons in hippocampal CA1 region of rats.Methods Wistar male rats(3—4 weeks)were killed by cervical dislocation and hippocampal slices(400 ?m)were prepared,blind whole-cell voltage-clamp recordings were performed on the CA1 pyramidal cells in hippocampal slices to examine and analyze the effects of PNS(0.05—0.4 g/L)on CA1 afferent fiber-evoked excitatory postsynaptic currents(EPSCs)and inhibitory postsynaptic currents(IPSCs),respectively.Moreover,the Schaffer collateral/commissural pathway was stimulated with paired pulses(interpulse interval was 50 ms)and the paired-pulse facilitation(PPF)was analyzed by EPSC2/EPSC1(P2/P1)ratio.Results PNS(0.1—0.4 g/L)significantly depressed amplitude of EPSCs in neurons in the hippocampal CA1 region(P0.05).Conclusion The inhibitory effect of PNS on EPSCs in hippocampal CA1 pyramidal neurons is not due to the reinforcement of the inhibiting interneurons.It may be a result of direct inhibition on excitatory synaptic transmission.The increasing of P2/P1 ratio after PNS application suggests that PNS depresses the excitatory synaptic transmission by presynaptic mechanism.