Integrity of perforant path fibers and the frequency of action potential independent excitatory and inhibitory synaptic events in dentate gyrus granule cells.

Whole-cell voltage clamp recordings in 400 microns thick hippocampal slices revealed discrete excitatory and inhibitory postsynaptic currents which persisted at synapses on granule cells following abolition of action potentials with 1 microM tetrodotoxin (TTX). The conductances associated with excitatory amino acid and GABAA receptor mediated events had mean peaks of 200 and 800 pS, and decayed monoexponentially with time constants of 5.6 and 5.3 ms. At a holding potential close to the normal resting membrane potential of granule cells (-80 to -90 mV), the frequency of glutamate/aspartate mediated spontaneous excitatory postsynaptic currents (sEPSCs) was decreased from 2.04 Hz in slices cut parallel to the plane of the perforant path to 0.87 Hz in slices cut in a plane that disrupted the distal perforant path fibres, suggesting that presynaptic integrity influences the rate of action potential independent neurotransmitter release. The orientation of the slicing had no effect on the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs).     

Intrinsic membrane properties determine hippocampal differential firing pattern in vivo in anesthetized rats

The hippocampus plays a key role in learning and memory. Previous studies suggested that the main types of principal neurons, dentate gyrus granule cells (GCs), CA3 pyramidal neurons, and CA1 pyramidal neurons, differ in their activity pattern, with sparse firing in GCs and more frequent firing in CA3 and CA1 pyramidal neurons. It has been assumed but never shown that such different activity may be caused by differential synaptic excitation. To test this hypothesis, we performed high‐resolution whole‐cell patch‐clamp recordings in anesthetized rats in vivo. In contrast to previous in vitro data, both CA3 and CA1 pyramidal neurons fired action potentials spontaneously, with a frequency of ～3–6 Hz, whereas GCs were silent. Furthermore, both CA3 and CA1 cells primarily fired in bursts. To determine the underlying mechanisms, we quantitatively assessed the frequency of spontaneous excitatory synaptic input, the passive membrane properties, and the active membrane characteristics. Surprisingly, GCs showed comparable synaptic excitation to CA3 and CA1 cells and the highest ratio of excitation versus hyperpolarizing inhibition. Thus, differential synaptic excitation is not responsible for differences in firing. Moreover, the three types of hippocampal neurons markedly differed in their passive properties. While GCs showed the most negative membrane potential, CA3 pyramidal neurons had the highest input resistance and the slowest membrane time constant. The three types of neurons also differed in the active membrane characteristics. GCs showed the highest action potential threshold, but displayed the largest gain of the input‐output curves. In conclusion, our results reveal that differential firing of the three main types of hippocampal principal neurons in vivo is not primarily caused by differences in the characteristics of the synaptic input, but by the distinct properties of synaptic integration and input‐output transformation. © 2015 The Authors Hippocampus Published by Wiley Periodicals, Inc.     

Ionic currents underlying spontaneous action potentials in isolated cerebellar Purkinje neurons.

Acutely dissociated cell bodies of mouse Purkinje neurons spontaneously fired action potentials at approximately 50 Hz (25 degrees C). To directly measure the ionic currents underlying spontaneous activity, we voltage-clamped the cells using prerecorded spontaneous action potentials (spike trains) as voltage commands and used ionic substitution and selective blockers to isolate individual currents. The largest current flowing during the interspike interval was tetrodotoxin-sensitive sodium current (approximately -50 pA between -65 and -60 mV). Although the neurons had large voltage-dependent calcium currents, the net current blocked by cobalt substitution for calcium was outward at all times during spike trains. Thus, the electrical effect of calcium current is apparently dominated by rapidly activated calcium-dependent potassium currents. Under current clamp, all cells continued firing spontaneously (though approximately 30% more slowly) after block of T-type calcium current by mibefradil, and most cells continued to fire after block of all calcium current by cobalt substitution. Although the neurons possessed hyperpolarization-activated cation current (Ih), little current flowed during spike trains, and block by 1 mM cesium had no effect on firing frequency. The outward potassium currents underlying the repolarization of the spikes were completely blocked by 1 mM TEA. These currents deactivated quickly (<1 msec) after each spike. We conclude that the spontaneous firing of Purkinje neuron cell bodies depends mainly on tetrodotoxin-sensitive sodium current flowing between spikes. The high firing rate is promoted by large potassium currents that repolarize the cell rapidly and deactivate quickly, thus preventing strong hyperpolarization and restoring a high input resistance for subsequent depolarization.     

Bidirectional pattern-specific plasticity of the slow afterhyperpolarization in rats: role for high-voltage activated Ca2+ channels and I h

A burst of action potentials in hippocampal neurons is followed by a slow afterhyperpolarization (sAHP) that serves to limit subsequent firing. A reduction in the sAHP accompanies acquisition of several types of learning, whereas increases in the sAHP are correlated with cognitive impairment. The present study demonstrates in vitro that activity-dependent bidirectional plasticity of the sAHP does not require synaptic activation, and depends on the pattern of action potential firing. Whole-cell current-clamp recordings from CA1 pyramidal neurons in hippocampal slices from young rats (postnatal days 14-24) were performed in blockers of synaptic transmission. The sAHP was evoked by action potential firing at gamma-related (50 Hz, gamma-AHP) or theta frequencies (5 Hz, theta-AHP), two firing frequencies implicated in attention and memory. Interestingly, when the gamma-AHP and theta-AHP were evoked in the same cell, a gradual potentiation of the gamma-AHP (186 ± 31%) was observed that was blocked using Ca(2+) channel blockers nimodipine (10 μm) or ω-conotoxin MVIIC (1 μm). In experiments that exclusively evoked the sAHP with 50 Hz firing, the gamma-AHP was similarly potentiated (198 ± 44%). However, theta-burst firing pattern alone resulted in a decrease (65 ± 19%) of the sAHP. In these experiments, application of the h-channel blocker ZD7288 (25 μm) selectively prevented enhancement of the gamma-AHP. These data demonstrate that induction requirements for bidirectional AHP plasticity depend on the pattern of action potential firing, and result from distinct mechanisms. The identification of novel mechanisms underlying AHP plasticity in vitro provides additional insight into the dynamic processes that may regulate neuronal excitability during learning in vivo.     

Active and Passive Membrane Properties of Spinal Cord Neurons that Are Rhythmically Active during Swimming in Xenopus Embryos

Cellular properties have been examined in ventrally located Xenopus spinal cord neurons that are rhythmically active during fictive swimming and presumed to be motoneurons. Resting potentials and input resistances of such neurons are - 75 +/- 2 mV (mean +/- standard error) and 118 +/- 17 M ohm respectively. Most cells fire a single impulse, 0.5 to 2.0 ms in duration and 48.5 +/- 1.8 mV in amplitude, in response to a depolarizing current step. A minority fire several spikes of diminishing amplitude to more strongly depolarizing current. Cells held above spike, threshold fire on rebound from brief hyperpolarizing pulses. Spikes are blocked by 0.1 to 1.0 microM tetrodotoxin (TTX) and are therefore Na+-dependent. Current/voltage (I/V) plots to injected current are approximately linear near the resting potential but become non-linear at more depolarized levels. Cells recorded in TTX with CsCI-filled microelectrodes show a linearized I/V plot at depolarized membrane potentials suggesting the normal presence of a voltage-dependent K+ conductance activated at relatively depolarized levels. Most cells recorded in this way but without TTX fire long trains of spikes of near constant amplitude, pointing to a role of the K+ conductance in limiting firing in normal cells. Spike blockage with TTX reveals, in some cells, a transient depolarizing Cd2+-sensitive and therefore presumably Ca2+-dependent potential that increases in amplitude with depolarization. Cells in TTX, Cd2+, and strychnine, and recorded with CsCI-filled microelectrodes to block active conductances respond to hyperpolarizing current steps with a two component exponential response. The cell time constant (tau0) obtained from the longer of these by exponential peeling is relatively long (mean 15.7 ms). These findings contribute to an increased understanding of the cellular properties involved in spinal rhythm generation in this simple vertebrate.     

NMDA receptors amplify mossy fiber synaptic inputs at frequencies up to at least 750 Hz in cerebellar granule cells

Neuronal integration of high‐frequency signals is important for rapid information processing. Cerebellar mossy fiber axons (MFs) can fire action potentials (APs) at frequencies of more than one kilohertz. However, it is unclear whether and how the postsynaptic cerebellar granule cells (GCs) are able to process these high‐frequency MF inputs. Here, we measured AP firing in GCs during high‐frequency MF stimulation and show that GC firing frequency increased non‐linearly when MF stimulation frequency was increased from 100 to 750 Hz. To investigate the mechanisms enabling such high‐frequency signaling, we analyzed the role of N‐methyl‐d ‐aspartate receptors (NMDARs), which have been implicated in synaptic signaling at lower frequencies. Application of D‐2‐amino‐5‐phosphonopentanoic acid (APV), a potent inhibitor of NMDARs, strongly impaired the GC firing frequency during high‐frequency MF stimulation. APV had no significant effect on single excitatory postsynaptic potentials (EPSPs) or currents (EPSCs) evoked at 1 Hz at resting membrane potentials. However, the time course of EPSCs evoked at 1 Hz at depolarized potentials or following high‐frequency MF stimulation was accelerated by APV. Thus, our results show that NMDAR‐mediated currents amplify high‐frequency MF inputs by prolonging the time courses of synaptic inputs, thereby causing greater synaptic summation of inputs. Hence, NMDARs support the integration of MF synaptic input at frequencies up to at least 750 Hz. Synapse 70:269–276, 2016. © 2016 Wiley Periodicals, Inc.     

Diversity in periodic pattern of firing in human hippocampal neurons

Firing periodicity was examined in human hippocampal neurons using autocorrelation analysis. Extracellular single-unit activities were recorded from the anterior hippocampus through fine platinum microelectrodes, and the typical firing pattern in an entire recording period was reconstructed statistically in autocorrelograms (average number of firings analyzed: 5639.0 +/- 968.1 SE, range: 1158 to 31,203; number of single-unit trains was 57). Three types of periodic firing were identified as highly consistent. The first pattern consisted of a random recurrence of high-frequency action potentials (100 to 300 Hz) and was observed as an intermittent burst. In this burst, the first 10 to 30 ms after the onset of the burst was the patterned firing of several action potentials, suggesting that the generation of this stereotyped portion of the burst is primarily due to intrinsic membrane characteristics. The second pattern was the continuous rhythmical firing with a lower frequency ranging from 1 to 30 Hz. The third pattern was a clustered rhythmical firing in which a series of short rhythmical firings recurred with regular intervals; the frequency of short rhythmical firing varied from 6.7 to 17 Hz between neurons, and the interval of the regular recurrence of these rhythmical firings ranged from 0.5 to 10 s among neurons. These firing periodicities not only cover a cellular rhythm in the theta frequency reported in the lower mammalian hippocampus but also appear to be more diverse than those previously reported for hippocampal neurons in the animal literature.     

神经肽Y对海马神经元“癫痫样”动作电位的影响

目的探讨神经肽Y(neuropeptide Y,NPY)对海马神经元"癫痫样"动作电位的影响。方法用无镁细胞外液处理原代培养12 d的海马神经元3 h,诱导海马神经元癫痫样放电,建立海马神经元癫痫样放电模型;用全细胞膜片钳电流钳模式检测神经元动作电位,分别给予0.1μmol/L和1μmol/L NPY各1μL,给药时间10 s,观察其对神经元动作电位频率及波幅的影响。结果无镁细胞外液处理神经元3 h,可以形成稳定的海马神经元癫痫样放电模型,频率16～23 Hz,波幅75～96 mV。模型组神经元动作电位频率为(18.00±2.32)Hz,而0.1μmol/L和1μmol/L NPY组分别为(4.75±1.04)Hz和(1.50±0.75)Hz。与模型组相比较,两种浓度NPY组均降低了动作电位发放的频率(P<0.05)。模型组神经元动作电位波幅为(82.25±5.17)mV,而0.1μmol/L和1μmol/L NPY组分别为(49.75±2.49)mV和(40.00±2.20)mV。与模型组相比较,两种浓度NPY组均降低了动作电位发放的波幅(P<0.05)。两种浓度NPY之间相比较,也有统计学差异(P<0.05)。1μmol/LNPY明显抑制了动作电位发放的频率和波幅。结论 NPY能够抑制无镁细胞外液诱发的神经元癫痫样电活动,为应用NPY抑制癫痫发作提供了细胞电生理学证据。     

Stimulus parameters affecting paired-pulse depression of dentate granule cell field potentials. II. Low-frequency stimulation

Low frequency (1 Hz) stimulation of the perforant path produces a depression in the population spike (PS) of dentate granule cell field potentials and also may affect the strength of paired pulse depression. The effects of 1 Hz stimulation (30 s train) on paired pulse depression (20 and 200 ms interpulse intervals, IPI) were evaluated in the unanesthetized rat under two conditions: (i) when the stimulus intensity of both pulses was increased simultaneously (5–100%); and (ii) when the stimulus intensity of the first (conditioning) pulse was increased (5–100%), while the stimulus intensity of the second (test) pulse was held constant (50%). The test PS amplitude was predicted based upon either the conditioning PS amplitude at the end of the 1 Hz train or upon the additive effects of paired pulse depression and 1 Hz stimulation. These predicted values then were assessed for the best fit to observed values following 1 Hz trains. Under both stimulus conditions, the 1 Hz depression in the conditioning PS amplitude exhibited characteristics that were identical to late paired pulse depression recorded before the train. A decrease in the test PS amplitude also was observed following 1 Hz stimulation at the 20 and 200 ms IPIs. The best fit to observed values of the test PS at the end of 1 Hz trains was provided by estimates based upon the additive effects of 1 Hz stimulation and paired pulse depression. These results indicate that the strenght of paired pulse depression in the unanesthetized rat is unchanged following 1 Hz stimulation, and further, that the 1 Hz depression in dentate granule cell field potentials most likely reflects the cumulative influence of late paired pulse depression.     

Enhanced Synaptic Inhibition Disrupts the Efferent Code of Cerebellar Purkinje Neurons in Leaner Cav2.1 Ca2+ Channel Mutant Mice

Cerebellar Purkinje cells (PCs) encode afferent information in the rate and temporal structure of their spike trains. Both spontaneous firing in these neurons and its modulation by synaptic inputs depend on Ca²⁺ current carried by Ca_v2.1 (P/Q) type channels. Previous studies have described how loss-of-function Ca_v2.1 mutations affect intrinsic excitability and excitatory transmission in PCs. This study examines the effects of the leaner mutation on fast GABAergic transmission and its modulation of spontaneous firing in PCs. The leaner mutation enhances spontaneous synaptic inhibition of PCs, leading to transitory reductions in PC firing rate and increased spike rate variability. Enhanced inhibition is paralleled by an increase in the frequency and amplitude of spontaneous inhibitory postsynaptic currents (sIPSCs) measured under voltage clamp. These differences are abolished by tetrodotoxin, implicating effects of the mutation on spike-induced GABA release. Elevated sIPSC frequency in leaner PCs is not accompanied by increased mean firing rate in molecular layer interneurons, but IPSCs evoked in PCs by direct stimulation of these neurons exhibit larger amplitude, slower decay rate, and a higher burst probability compared to wild-type PCs. Ca²⁺ release from internal stores appears to be required for enhanced inhibition since differences in sIPSC frequency and amplitude in leaner and wild-type PCs are abolished by thapsigargin, an ER Ca²⁺ pump inhibitor. These findings represent the first account of the functional consequences of a loss-of-function P/Q channel mutation on PC firing properties through altered GABAergic transmission. Gain in synaptic inhibition shown here would compromise the fidelity of information coding in these neurons and may contribute to impaired cerebellar function resulting from loss-of function mutations in the Ca_V2.1 channel gene.     

Synchronization of GABAergic interneuronal networks during seizure-like activity in the rat horizontal hippocampal slice

We studied the contribution of GABAergic (gamma-aminobutyric acid) neurotransmission to epileptiform activity using the horizontal hippocampal rat brain slice. Seizure-like (ictal) activity was evoked in the CA1 area by applying high-frequency trains (80 Hz for 2 s) to the Schaffer collaterals. Whole-cell recordings from stratum oriens-alveus interneurons revealed burst firing with superimposed high-frequency spiking which was synchronous with field events and pyramidal cell firing during ictal activity. On the other hand, interictal interneuronal bursts were synchronous with large-amplitude inhibitory postsynaptic potentials (IPSPs) in pyramidal cells. Excitatory and inhibitory postsynaptic potentials were simultaneously received by pyramidal neurons during the ictal afterdischarge, and were synchronous with interneuronal bursting and field potential ictal events. The GABAA receptor antagonist bicuculline greatly reduced the duration of the ictal activity in the CA1 layer, and evoked rhythmic interictal synchronous bursting of interneurons and pyramidal cells. With intact GABAergic transmission, interictal field potential events were synchronous with large amplitude IPSPs (9.8 +/- 2.4 mV) in CA1 pyramidal cells, and with interneuronal bursting. Simultaneous dual recordings revealed synchronous IPSPs received by widely separated pyramidal neurons during ictal and interictal periods, indicative of widespread interneuronal firing synchrony throughout the hippocampus. CA3 pyramidal neurons fired in synchrony with interictal field potential events recorded in the CA1 layer, and glutamate receptor antagonists abolished interictal interneuronal firing and synchronous large amplitude IPSPs received by CA1 pyramidal cells. These observations provide evidence that the interneuronal network may be entrained in hyperexcitable states by GABAergic and glutamatergic mechanisms.     

Neurogliaform cells in the molecular layer of the dentate gyrus as feed-forward γ-aminobutyric acidergic modulators of entorhinal-hippocampal interplay

Feed-forward inhibition from molecular layer interneurons onto granule cells (GCs) in the dentate gyrus is thought to have major effects regulating entorhinal-hippocampal interactions, but the precise identity, properties, and functional connectivity of the GABAergic cells in the molecular layer are not well understood. We used single and paired intracellular patch clamp recordings from post-hoc-identified cells in acute rat hippocampal slices and identified a subpopulation of molecular layer interneurons that expressed immunocytochemical markers present in members of the neurogliaform cell (NGFC) class. Single NGFCs displayed small dendritic trees, and their characteristically dense axonal arborizations covered significant portions of the outer and middle one-thirds of the molecular layer, with frequent axonal projections across the fissure into the CA1 and subicular regions. Typical NGFCs exhibited a late firing pattern with a ramp in membrane potential prior to firing action potentials, and single spikes in NGFCs evoked biphasic, prolonged GABA(A) and GABA(B) postsynaptic responses in GCs. In addition to providing dendritic GABAergic inputs to GCs, NGFCs also formed chemical synapses and gap junctions with various molecular layer interneurons, including other NGFCs. NGFCs received low-frequency spontaneous synaptic events, and stimulation of perforant path fibers revealed direct, facilitating synaptic inputs from the entorhinal cortex. Taken together, these results indicate that NGFCs form an integral part of the local molecular layer microcircuitry generating feed-forward inhibition and provide a direct GABAergic pathway linking the dentate gyrus to the CA1 and subicular regions through the hippocampal fissure.     

Calbindin-D28k content and firing pattern of hippocampal granule cells in amygdala-kindled rats: a perforated patch-clamp study

The dentate gyrus is believed to play an important pathophysiological role during experimentally induced kindling. In this study, we investigated whether an altered content of the calcium binding protein calbindin-D(28k) or an increased intrinsic excitability of hippocampal granule cells contribute to the induction of the kindling phenomenon. We determined the firing pattern of granule cells in hippocampal slices using perforated patch-clamp recordings in current clamp mode. The expression of calbindin-D(28k) and glutamic acid decarboxylase (GAD(67)) by granule cells was analyzed immunohistochemically. Rats developed secondarily generalized limbic seizures within approximately 11 days of twice-daily stimulation of the amygdala. As reported for other kindling paradigms, this protocol induced a clear up-regulation of GAD(67) in granule cells, indicating their involvement in the induced neuronal activity. However, when comparing kindled and control rats, we could not detect any differences in intrinsic excitability: Firing frequency, after-hyperpolarisations, action potentials, input resistance and membrane potentials were nearly identical between both groups. Furthermore, we did not observe any differences in the calbindin-D(28k) immunoreactivity between groups. In every slice, virtually all granule cells were found to be strongly calbindin-D(28k) positive, and there was no apparent reduction in the general level of calbindin-D(28k) expression. We conclude that changes in intrinsic membrane properties or in the calbindin-D(28k) content of granule cells are not necessary for the development of amygdala kindling.     

In vivo intracellular correlates of hippocampal formation theta-on and theta-off cells.

Using urethane-anesthetized rat, intracellular recordings were made in hippocampal formation cells classified according to previously established criteria as either theta-on or theta-off, in order to further define the electrophysiological characteristics of these cells. Four cells classified as phasic theta-off cells had short duration spikes (less than 1 ms), high input resistances (54-61 M omega) and large fast afterhyperpolarizations (6-10 mV), thus sharing some of the properties of identified hippocampal interneurons. Phasic theta-off cells also exhibited rhythmic membrane potential oscillations (MPOs) ranging from 4 to 10 mV in amplitude during the simultaneous occurrence of extracellular theta field activity, but not during the occurrence of large amplitude irregular field activity (LIA). The MPOs of phasic theta-off cells were the same frequency as and were highly coherent with the extracellular theta field activity. In all four phasic theta-off cells the positive peak of the MPO was in phase with the positive peak of the local theta field activity. At the onset of extracellular theta field activity above 4-5 Hz, the membrane potentials of phasic theta-off cells showed a 5-10-mV hyperpolarizing shift, accompanied by MPOs without spike discharges. As theta frequency slowed down there was a return to baseline membrane potential levels and spike discharges occurred near the positive peak of the MPOs. The seven cells classified as phasic theta-on had longer duration spikes (greater than 1 ms), lower input resistances (22-36 M omega) and small (approx. 1.0 mV) fast afterhyperpolarizations, thus sharing some of the properties of hippocampal projection cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Sodium conductance in cultured myenteric AH-type neurons from guinea-pig small intestine

Whole-cell patch clamp methods were used to investigate sodium conductance in after-hyperpolarization-type (AH) enteric neurons in culture after dissociation from the myenteric plexus of guinea-pig small intestine. Inward current carried by Na+ (I(Na)) was identified and its current-voltage characteristics were compared with those for inward Ca2+ current (I(Ca)). The I(Na) current was a rapidly inactivating current relative to I(Ca). Application of tetrodotoxin (TTX) blocked I(Na) with an EC50 of 10.7 nM. Activation curves for I(Na) showed a rapid decrease in time to peak for test potentials from holding potentials of -80 mV to between -40 and -10 mV. Voltage-dependence of steady-state inactivation curves for I(Na) was fit to the Boltzmann equation with potential for half-inactivation (V(1/2)) = -55.6 mV and slope factor (k) = 6.4 mV. Steady-state inactivation for I(Ca) fit the Boltzmann equation with a V(1/2) = -38.9 mV and k= 14.4 mV. Kinetics for inactivation of I(Na) were voltage dependent at potentials between -70 and -30 mV and accelerated and became less voltage-dependent at more positive potentials. The time constant (tau) for inactivation at -70 mV was tau = 161 +/- 23 ms and decreased to tau = 2.3 +/- 0.2 ms at -30 mV. Rapid acceleration of inactivation occurred between -50 and -40 mV. This was also the range where activation began. Recovery from inactivation with the membrane potential clamped at -100 or -80 mV was rapid and fit by a single exponential with tau = 7.3 +/- 1.1 ms for -100 mV and 21.5 +/- 5.1 ms for -80 mV. The results suggest that AH-type enteric neurons have only one type of Na+ channel that behaves like the "classical" voltage-gated tetrodotoxin-sensitive fast channel. The findings support the hypothesis that I(Na) current is an important factor in determination of excitability and firing behavior in AH neurons. I(Na) and I(Ca) together determine the properties of the rising phase of the spike and thereby contribute to global determinants of excitability as the neurons are exposed to multiple depolarizing and hyperpolarizing stimuli from synaptic inputs and mediators released from enteroparacrine cells.     

Morphological and physiological properties of rat cerebellar neurons in mature and developing cultures

Immunohistochemical techniques and antibodies to gamma-aminobutyric acid (GABA), parvalbumin, and cyclic guanosine monophosphate-dependent protein kinase were used to identify populations of cerebellar neurons in culture that exhibit morphological features and immunoreactivity characteristic of neuronal types present in the cortical region of the cerebellum in vivo. The cultures were examined at 3 culture ages: 6-9, 12-15 and greater than 15 days in vitro, reflecting early, intermediate and late periods in cerebellar development. Neurons identified as Purkinje neurons (PNs), granule cells or inhibitory interneurons (stellate, basket, Golgi and Lugaro cells) were present at all culture ages. The granule cells (GCs) and inhibitory interneurons (INs) were morphologically well developed at the youngest culture age studied; morphological features did not change dramatically during the culture period. In contrast, the PNs were morphologically immature at 6-9 DIV (DIV = days in vitro) and exhibited dramatic changes in morphological structure with culture age. Extracellular recordings from PNs. GCs and INs in living cultures revealed that all classes of neurons exhibited spontaneous activity, but that only a portion of the GCs and INs were spontaneously active. The spontaneously active GCs and INs exhibited variable patterns of activity and low firing rates (approximately 2-6 Hz) at all culture ages studied. At 6 DIV, PNs exhibited firing rates and patterns similar to that of the interneurons. At older culture ages, the firing rate and pattern of PNs was significantly different from the GCs and INs and was characterized by high frequency (greater than 10 Hz) spike activity usually in a regular pattern. All cerebellar neurons by excited by the transmitter glutamate (Glu). The Glu response in the GCs and INs consisted of a brief burst of single spikes; in PNs, the response to Glu was prolonged and multiphasic. These data indicate that the cerebellar GCs and INs express morphological, physiological and developmental properties that are significantly different from the PN.     

Properties of synaptic currents during non-adrenergic inhibition of the smooth muscle cells of the large intestine in the guinea pig

Inhibitory junctional currents (IJCs) were recorded under voltage clamp conditions in response to brief transmural stimulation of the circular muscle of the guinea pig colon using the double sucrose gap method in the presence of atropine. The time course of IJC decay was approximately exponential 100-150 ms after the peak value. The IJC amplitude depended linearly on the membrane potential with the reversal potential (-70 mV) near the potassium equilibrium potential. The time constant (tau) of the IJC decay depended exponentially on the membrane potential and became e-fold decreased when the membrane was hyperpolarized approximately by 120 mV. Varying the quantal content of IJC caused an increase of tau upon rising the amount or transmitter released and its decrease with the depression of IJC. Application of ATP (10(-3)M) caused a decrease of tau and IJC amplitude, while apamine reduced the amplitude of IJC without any changes in their time course. The results are discussed in terms of a buffered diffusion hypothesis supposing a cooperative action of transmitter released on junctional receptors.     

Electrical properties and synaptic potentials of rabbit pancreatic neurons

Pancreatic ganglia receive innervation from a wide variety of extrinsic nerves and supply the predominant innervation to pancreatic acini, islets, and ducts. This study used intracellular recordings to investigate the electrical properties and synaptic potentials of rabbit pancreatic neurons. Neurons had a mean resting membrane potential of -54+/-0.4 mV and generated action potentials with a mean overshoot of 10+/-0.4 mV and a mean after-spike hyperpolarization (ASH) of 11+/-0.5 mV with duration of 210+/-19 ms. Action potentials exhibited a high threshold (-15+/-1 mV) for intracellular stimulation and a phasic firing pattern was observed in response to prolonged depolarizing currents. Stimulation of attached nerve bundles evoked multiple fast excitatory postsynaptic potentials (fEPSPs) which were abolished by hexamethonium in 75% of neurons, while a non-cholinergic fEPSP was observed in 25% of the neurons. Repetitive stimulation (3-30 Hz) evoked muscarinic slow EPSPs with a mean amplitude of 8+/-2 mV and duration of 5+/-1 s in a small subset (21%) of neurons. Exogenous muscarine evoked a mean slow depolarization of 10+/-1 mV amplitude in 22% of neurons tested. Following repetitive nerve stimulation non-cholinergic late, slow EPSPs with a mean amplitude of 4.3+/-0.4 mV were recorded in 32% of neurons. Nicotinic transmission was subject to inhibition mediated by presynaptic muscarinic receptors at low (0.5 Hz) stimulus frequencies in 80% of neurons. At higher frequencies (> or =1 Hz), either facilitation or depression of nicotinic transmission was observed depending on the ganglion studied. A population (9%) of neurons exhibited spontaneous, low-amplitude pacemaker-like potentials. Spontaneous fEPSPs and action potentials were also observed and these occasionally occurred in rhythmically timed bursts. Thus, distinct subpopulations of pancreatic neurons could be identified on the basis of both their intrinsic electrical properties and the receptors mediating and/or modulating synaptic transmission. These neurons function as critical sites of integration for synaptic input from extrinsic pancreatic nerves and thereby determine the postganglionic firing patterns presented to the pancreatic exocrine and endocrine secretory cells.     

Feedforward inhibition is randomly wired from individual granule cells onto CA3 pyramidal cells

Feedforward inhibition (FFI) between the dentate gyrus (DG) and CA3 sparsifies and shapes memory‐ and spatial navigation‐related activities. However, our understanding of this prototypical FFI circuit lacks essential details, as the wiring of FFI is not yet mapped between individual DG granule cells (GCs) and CA3 pyramidal cells (PCs). Importantly, theoretically opposite network contributions are possible depending on whether the directly excited PCs are differently inhibited than the non‐excited PCs. Therefore, to better understand FFI wiring schemes, we compared the prevalence of disynaptic inhibitory postsynaptic events (diIPSCs) between pairs of individually recorded GC axons or somas and PCs, some of which were connected by monosynaptic excitation, while others were not. If FFI wiring is specific, diIPSCs are expected only in connected PCs; whereas diIPSCs should not be present in these PCs if FFI is laterally wired from individual GCs. However, we found single GC‐elicited diIPSCs with similar probabilities irrespective of the presence of monosynaptic excitation. This observation suggests that the wiring of FFI between individual GCs and PCs is independent of the direct excitation. Therefore, the randomly distributed FFI contributes to the hippocampal signal sparsification by setting the general excitability of the CA3 depending on the overall activity of GCs.     

A transient voltage-dependent outward potassium current in mammalian cerebellar Purkinje cells

Utilizing the single electrode voltage clamp technique applied to rat Purkinje cells (PCs), we have recorded a transient outward voltage-dependent potassium current, I_to. Half maximal values for inactivation and activation were −65 mV and −39 mV, respectively. I_to decays as a single exponential with a time constant of 95 ms and is reduced by 4-aminopyridine. Based on criteria of voltage dependency of activation and inactivation, kinetics of inactivation, ionic selectivity and pharmacologic sensitivity, we have verified a strong resemblance between the typical A current in other neurons, and I_to in PCs. As in other cells, I_to may be important in shaping PC output by modulating intrinsic and extrinsic factors that govern PC firing.