Experiments were performed to study the voltage-dependence of miniature inhibitory postsynaptic current (mIPSC) frequency and amplitude using patch-clamp technique with whole cell recording in optic tectal slices of Xenopus. The following results have been observed. (1) When the membrane potentials of a neuron were depolarized or hyperpolarized stepwise from a resting potential via recording pipette to inject a DC current, the frequency and/or amplitude of mIPSCs increased or decreased respectively. The frequency of mIPSCs increased gradually with depolarizing membrane potential and it attained to the maximum as the membrane potential was held at +10 mV. (2) The amplitude increased slightly as the neuron was depolarized. When the depolarization of membrane potential reached -30 or -40 mV, the amplitudes of mIPSCs were maximal. Further depolarization resulted in a decrease of amplitude. Meanwhile, the large mIPSCs appeared when the membrane potential depolarized to a range between -20 mV and +10 mV. (3) With Ca(2+)-free bath solution, the frequency and amplitude of mIPSCs also increased stepwise progressively on depolarization of membrane potential, but the increase was less marked as corresponding value in normal saline perfusion. (4) When the [K(+)](o) in bath solution increased, the frequency of mIPSCs decreased markedly and the amplitude of mIPSCs decreased slightly. If the external K(+) concentration increased further to higher than 20 mmol/L, the neuron produced a marked slow inward or outward membrane current. The possible mechanism underlying the voltage-dependence of mIPSC frequency and amplitude is discussed briefly.
Animals ; Brain ; cytology ; physiology ; Inhibitory Postsynaptic Potentials ; physiology ; Membrane Potentials ; physiology ; Miniature Postsynaptic Potentials ; physiology ; Neurons ; physiology ; Patch-Clamp Techniques ; Potassium Channels, Voltage-Gated ; physiology ; Xenopus

OBJECTIVETo observe the microstructure of the cell membrane of epileptic neurons using atomic force microscopy (AFM).

METHODSModel of epileptic neurons was established by subjecting the neurons culture for 14 days in vitro to magnesium-free media treatment for 3 h. Patch clamp technique was applied to record the electrophysiological activity of the epileptic neurons. AFM was performed to observe and measure the microstructure of the cell membrane of the epileptic neuron.

RESULTSAfter a 3-hour treatment with magnesium-free media, the epileptic neurons displayed sustained epileptiform discharge, which continued after the neurons were returned to normal medium culture on day 14. Under AFM scanning size of 80 microm x 80 microm and 2 microm x 2 microm, no obvious difference in the morphology of the cell membrane was noted between epileptic and normal neurons; under the scanning size of 500 nm x 500 nm, small pits occurred in the cell membrane in both groups, but no significant difference was found in the dimension of the pits between the two groups (the diameter and depth of the pits was 114.86-/+9.33 nm and 5.71-/+0.69 nm in epileptic neurons, and 116.4-/+9.13 nm and 5.69-/+0.71 nm in the control neurons, respectively, P>0.05).

CONCLUSIONAFM provides a new method for observing neuronal membrane microstructure at nanometer resolutions. No significant alterations occur in the membrane of the neurons after a 3-hour magnesium-free media treatment.

Cell Membrane ; ultrastructure ; Cells, Cultured ; Culture Media ; Epilepsy ; pathology ; Excitatory Postsynaptic Potentials ; Inhibitory Postsynaptic Potentials ; Magnesium ; Microscopy, Atomic Force ; Neurons ; ultrastructure ; Patch-Clamp Techniques

The hypothalamic paraventricular nucleus (PVN) contains two types of neurons projecting to either the rostral ventrolateral medulla (PVN(RVLM)) or the intermediolateral horn (IML) of the spinal cord (PVN(IML)). These two neuron groups are intermingled in the same subdivisions of the PVN and differentially regulate sympathetic outflow. However, electrophysiological evidence supporting such functional differences is largely lacking. Herein, we compared the electrophysiological properties of these neurons by using patch-clamp and retrograde-tracing techniques. Most neurons (>70%) in both groups spontaneously fired in the cell-attached mode. When compared to the PVN(IML) neurons, the PVN(RVLM) neurons had a lower firing rate and a more irregular firing pattern (p < 0.05). The PVN(RVLM) neurons showed smaller resting membrane potential, slower rise and decay times, and greater duration of spontaneous action potentials (p < 0.05). The PVN(RVLM) neurons received greater inhibitory synaptic inputs (frequency, p < 0.05) with a shorter rise time (p < 0.05). Taken together, the results indicate that the two pre-sympathetic neurons differ in their intrinsic and extrinsic electrophysiological properties, which may explain the lower firing activity of the PVN(RVLM) neurons. The greater inhibitory synaptic inputs to the PVN(RVLM) neurons also imply that these neurons have more integrative roles in regulation of sympathetic activity.
Action Potentials ; Animals ; Fires ; Horns ; Inhibitory Postsynaptic Potentials ; Membrane Potentials ; Neurons ; Paraventricular Hypothalamic Nucleus ; Patch-Clamp Techniques ; Spinal Cord ; Spinal Cord Lateral Horn

PURPOSE: Modafinil is a wake-promoting agent that has been proposed to improve cognitive performance at the preclinical and clinical levels. Since there is insufficient evidence for modafinil to be regarded as a cognitive enhancer, the aim of this study was to investigate the effects of chronic modafinil administration on behavioral learning in healthy adult rats. METHODS: Y-maze training was used to assess learning performance, and the whole-cell patch clamp technique was used to assess synaptic transmission in pyramidal neurons of the hippocampal CA1 region of rats. RESULTS: Intraperitoneal administration of modafinil at 200 mg/kg or 300 mg/kg significantly improved learning performance. Furthermore, perfusion with 1mM modafinil enhanced the frequency and amplitude of spontaneous postsynaptic currents and spontaneous excitatory postsynaptic currents in CA1 pyramidal neurons in hippocampal slices. However, the frequency and amplitude of spontaneous inhibitory postsynaptic currents in CA1 pyramidal neurons were inhibited by treatment with 1mM modafinil. CONCLUSIONS: These results indicate that modafinil improves learning and memory in rats possibly by enhancing glutamatergic excitatory synaptic transmission and inhibiting GABAergic (gamma-aminobutyric acid-ergic) inhibitory synaptic transmission.
Adult ; Animals ; CA1 Region, Hippocampal ; Excitatory Postsynaptic Potentials ; Humans ; Inhibitory Postsynaptic Potentials ; Learning* ; Memory ; Neurons ; Perfusion ; Rats* ; Synaptic Potentials ; Synaptic Transmission*

Reactive oxygen species (ROS), which include hydrogen peroxide (H2O2), the superoxide anion (O2-.), and the hydroxyl radical (OH.), are generated as by-products of oxidative metabolism in cells. The cerebral cortex has been found to be particularly vulnerable to production of ROS associated with conditions such as ischemia-reperfusion, Parkinson's disease, and aging. To investigate the effect of ROS on inhibitory GABAergic synaptic transmission, we examined the electrophysiological mechanisms of the modulatory effect of H2O2 on GABAergic miniature inhibitory postsynaptic current (mIPSCs) in mechanically isolated rat cerebral cortical neurons retaining intact synaptic boutons. The membrane potential was voltage-clamped at -60 mV and mIPSCs were recorded and analyzed. Superfusion of 1-mM H2O2 gradually potentiated mIPSCs. This potentiating effect of H2O2 was blocked by the pretreatment with either 10,000-unit/mL catalase or 300-micrometer N-acetyl-cysteine. The potentiating effect of H2O2 was occluded by an adenylate cyclase activator, forskolin, and was blocked by a protein kinase A inhibitor, N-(2-[p-bromocinnamylamino] ethyl)-5-isoquinolinesulfonamide hydrochloride. This study indicates that oxidative stress may potentiate presynaptic GABA release through the mechanism of cAMP-dependent protein kinase A (PKA)-dependent pathways, which may result in the inhibition of the cerebral cortex neuronal activity.
Adenylyl Cyclases ; Aging ; Animals ; Catalase ; Cerebral Cortex ; Cyclic AMP-Dependent Protein Kinases ; Forskolin ; gamma-Aminobutyric Acid ; Hydrogen Peroxide ; Hydroxyl Radical ; Inhibitory Postsynaptic Potentials ; Membrane Potentials ; Neurons ; Oxidative Stress ; Parkinson Disease ; Presynaptic Terminals ; Rats ; Reactive Oxygen Species ; Superoxides ; Synaptic Transmission

AIMTo investigate the effects of morphine on synaptic transmission of neurons of central nervous system and reveal the mechanism underlying it.

METHODSNew born wistar rats were used for primary culture of hippocampus neurons. Using whole-cell patch-clamp technique, we observed the excitatory and spontaneous inhibitory postsynaptic current (EPSC, sIPSC) and glutamate-induced current before and after morphine treatment.

RESULTS(1) sEPSC of hippocampal neurons was markedly increased after morphine application. The effect of morphine was blocked by opioid antagonist naloxone (n=18, P < 0.01). (2) The frequency of mEPSC and the amplitude of glutamate-induced current of hippocampal neurons had no significant changes after morphine treatment (P > 0.05). (3) Morphine inhibited sIPSC of hippocampal neurons markedly and naloxone could block this effect (n=13, P < 0.01).

CONCLUSIONThe results suggest that the exciting effect of morphine on hippocampal neurons are not due to direct influence of morphine on glutamate synapses transmission, but may result from the inhibition on interneurons, that is "disinhibition" way.

Animals ; Animals, Newborn ; Cells, Cultured ; Excitatory Postsynaptic Potentials ; physiology ; Hippocampus ; cytology ; Inhibitory Postsynaptic Potentials ; Morphine ; pharmacology ; Neurons ; drug effects ; physiology ; Patch-Clamp Techniques ; Rats ; Rats, Wistar ; Synaptic Transmission ; drug effects ; physiology

The medial vestibular nucleus (MVN) neurons are controlled by excitatory synaptic transmission from the vestibular afferent and commissural projections, and by inhibitory transmission from interneurons. Spontaneous synaptic currents of MVN neurons were studied using whole cell patch clamp recording in slices prepared from 13- to 17-day-old rats. The spontaneous inhibitory postsynaptic currents (sIPSCs) were significantly reduced by the GABAA antagonist bicuculline (20micrometer), but were not affected by the glycine antagonist strychnine (1micrometer). The frequency, amplitude, and decay time constant of sIPSCs were 4.3 0.9 Hz, 18.1 2.0 pA, and 8.9 0.4 ms, respectively. Spontaneous excitatory postsynaptic currents (sEPSCs) were mediated by non-NMDA and NMDA receptors. The specific AMPA receptor antagonist GYKI-52466 (50micrometer) completely blocked the non-NMDA mediated sEPSCs, indicating that they are mediated by an AMPA-preferring receptor. The AMPA mediated sEPSCs were characterized by low frequency (1.5 0.4 Hz), small amplitude (13.9 1.9 pA), and rapid decay kinetics (2.8 0.2 ms). The majority (15/21) displayed linear I-V relationships, suggesting the presence of GluR2-containing AMPA receptors. Only 35% of recorded MVN neurons showed NMDA mediated currents, which were characterized by small amplitude and low frequency. These results suggest that the MVN neurons receive excitatory inputs mediated by AMPA, but not kainate, and NMDA receptors, and inhibitory transmission mediated by GABAA receptors in neonatal rats.
alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid ; Animals ; Bicuculline ; Excitatory Postsynaptic Potentials* ; Glycine ; Inhibitory Postsynaptic Potentials ; Interneurons ; Kainic Acid ; Kinetics ; N-Methylaspartate ; Neurons* ; Rats* ; Receptors, AMPA ; Receptors, N-Methyl-D-Aspartate ; Strychnine ; Synaptic Transmission ; Vestibular Nuclei*

Following peripheral nerve injury, excessive nociceptive inputs result in diverse physiological alterations in the spinal cord substantia gelatinosa (SG), lamina II of the dorsal horn. Here, I report the alterations of excitatory or inhibitory transmission in the SG of a rat model for neuropathic pain ("spared nerve injury"). Results from whole-cell recordings of SG neurons show that the number of distinct primary afferent fibers, identified by graded intensity of stimulation, is increased at 2 weeks after spared nerve injury. In addition, short-term depression, recognized by paired-pulse ratio of excitatory postsynaptic currents, is significantly increased, indicating the increase of glutamate release probability at primary afferent terminals. The peripheral nerve injury also increases the amplitude, but not the frequency, of spontaneous inhibitory postsynaptic currents. These data support the hypothesis that peripheral nerve injury modifies spinal pain conduction and modulation systems to develop neuropathic pain.
Animals ; Depression ; Excitatory Postsynaptic Potentials ; Glutamic Acid ; Horns ; Inhibitory Postsynaptic Potentials ; Models, Animal ; Neuralgia ; Neurons ; Patch-Clamp Techniques ; Peripheral Nerve Injuries* ; Peripheral Nerves* ; Rats* ; Spinal Cord* ; Substantia Gelatinosa* ; Synaptic Transmission*

This study used in vivo intracellular recording in rat hippocampus to evaluate the effect of lidocaine and MK-801 on the membrane properties and the synaptic responses of individual neurons to electrical stimulation of the commissural pathway. Cells in control group typically fired in a tonic discharge mode with an average firing frequency of 2.4+/-0.9 Hz. Neuron in MK-801 treated group (0.2 mg/kg, i.p.) had an average input resistance of 32.8 +/- 5.7 Mg and a membrane time constant of 7.4 +/- 1. 8 ms. These neurons exhibited 2.4 +/- 0.2 ms spike durations, which were similar to the average spike duration recorded in the neurons of the control group. Slightly less than half of these neurons were firing spontaneously with an average discharge rate of 2.4 +/- 1.1 Hz. The average peak amplitude of the ABP following the spikes in these groups was 7.4+/-0.6 mV with respect to the resting membrane potentiaL Cells in MK-801 and lidocaine treated group (5 mg/kg, i.c.v.) had an average input resistance of 34.5+/-6.0 Mg and an average time constant of 8.0+/-1.4 ms. The cells were firing spontaneously at an average discharge rate of 0.6+/-0.4 Hz. Upon depolarization of the membrane by 0.8 nA for 400 ms, all of the tested cells exhibited accommodation of spike discharge. The most common synaptic response contained an EPSP followed by early-IPSP and late-IPSP. Analysis of the voltage dependence revealed that the early-IPSP and late-IPSP were putative Cl-and K+-dependent, respectively. Systemic injection of the NMDA receptor blocker, MK-801, did not block synaptic responses to the stimulation of the commissural pathway. No significant modifications of EPSP, early-IPSP, or late-IPSP components were detected in the MK-801 and/or lidocaine treated group. These results suggest that MK-801 and lidocaine manifest their CNS effects through firing pattern of hippocampal pyramidal cells and neural network pattern by changing the synaptic efficacy and cellular membrane properties.
Anesthetics, Local ; Animals ; Dizocilpine Maleate* ; Electric Stimulation ; Excitatory Postsynaptic Potentials ; Fires ; Hippocampus ; Lidocaine* ; Membrane Potentials ; Membranes ; N-Methylaspartate ; Neurons* ; Pyramidal Cells ; Rats

BACKGROUND AND OBJECTIVES: We are developing an in vitro preparation of the mouse inner ear so as to study morphophysiologic character of primary vestibular afferents and synaptic transmission within the vestibular epithelium. MATERIALS AND METHOD: We have intra-axonally recorded from over 300 ampullary fibers, close to the base of their respective anterior and lateral crista (<500 micrometer from hair cell/afferent nerve synapse), and labelled as a sub-set of these with biocytin (n=71). Discharge activity can be classified as regular or irregular based on the variation of the interspike interval (coefficient of variation). Using a micropusher to indent exposed windows of membranous labyrinth, we have characterized the response properties of both anterior and horizontal canal afferents. We studied afferent activity in response to sinusoidal indentations of anterior and horizontal membranous canal. RESULTS: The majority of labelled units were dimorphic (56 out of 71), having both calyx and bouton terminals and there was no labelled bouton terminal. Whether action potentials (Aps) were spontaneous or elicited with current, a heterogeneity of discharge activity was observed and these were similar to those previously reported in in vivo recordings from other mammalian species. In recordings over a range of frequencies from 0.01 to 10.0Hz, afferents responded with sinusoidal changes at discharge rates and modulation of membrane potential in a predictable manner. The phase response of the afferent discharge was characterized by frequency-dependent shifts in peak activity. The peak activity of anterior canal was in advance of the maximum indentation (180dgrees out of phase), with largest phase leads at 0.01 Hz (59.2+/-14.1dgrees) and the smallest phase leads occurring at 1.0 Hz (13.4+/-9.3dgrees), while maximum indentation was in advance of the peak activity at 10.0 Hz (-17.6+/-9.1dgrees). These phase shifts were similar to those reported in in vivo recordings from mammals, despite our use of artificial rather than natural rotational stimuli. CONCLUSION: We developed an in-vitro mouse model to study morphophysiologic characteristics of primary vestibular afferent nerve and synaptic transmission.
Action Potentials ; Animals ; Ear, Inner* ; Electrophysiology ; Epithelium ; Hair ; Mammals ; Membrane Potentials ; Mice* ; Population Characteristics ; Semicircular Canals ; Synaptic Transmission