Regulation of synaptic input to hypothalamic presympathetic neurons by GABA(B) receptors

The hypothalamic paraventricular (PVN) neurons projecting to the spinal cord and brainstem play an important role in the control of homeostasis and the sympathetic nervous system. Although GABA(B) receptors are present in the PVN, their function in the control of synaptic inputs to PVN presympathetic neurons is not clear. Using retrograde tracing and whole-cell patch-clamp recordings in rat brain slices, we determined the role of presynaptic GABA(B) receptors in regulation of glutamatergic and GABAergic inputs to spinally projecting PVN neurons. The GABA(B) receptor agonist baclofen (1-50 microM) dose-dependently decreased the frequency but not the amplitude of spontaneous excitatory postsynaptic currents (sEPSCs) and inhibitory postsynaptic currents (sIPSCs). The effect of baclofen on sEPSCs and sIPSCs was completely blocked by 10 microM CGP52432, a selective GABA(B) receptor antagonist. Baclofen also significantly reduced the frequency of both miniature excitatory and miniature inhibitory postsynaptic currents (mEPSCs and mIPSCs). Furthermore, uncoupling pertussis toxin-sensitive G(i/o) proteins with N-ethylmaleimide abolished baclofen-induced inhibition of mEPSCs and mIPSCs. However, the inhibitory effect of baclofen on the frequency of mIPSCs and mEPSCs persisted in the presence of either Cd2+, a voltage-gated Ca2+ channel blocker, or 4-aminopyridine, a blocker of voltage-gated K+ channels. Our results suggest that activation of presynaptic GABA(B) receptors inhibits synaptic GABA and glutamate release to PVN presympathetic neurons. This presynaptic action of GABA(B) receptors is mediated by the N-ethylmaleimide-sensitive G(i/o) proteins, but independent of voltage-gated Ca2+ and K+ channels.     

Morphological characteristics and central projections of two types of interneurons in the visual pathway of Hermissenda.

The synaptic interactions between photoreceptors in the eye and second-order neurons in the optic ganglion of the nudibranch mollusk Hermissenda are well characterized. However, the higher-order neural circuitry of the visual system, consisting of cerebropleural interneurons that receive synaptic input from photoreceptors and project to pedal motor neurons that mediate visually guided behaviors, is only partially understood. In this report we have examined the central projections of two identified classes of cerebropleural interneurons that receive excitatory or inhibitory synaptic input from identified photoreceptors. The classification of the interneurons was based on both morphological and electrophysiological criteria. Type I interneurons received monosynaptic excitatory or inhibitory synaptic input from identified photoreceptors and projected to postsynaptic targets within the cerebropleural ganglion. Type II interneurons, characterized here for the first time, received polysynaptic excitatory or inhibitory synaptic input from identified photoreceptors and projected to postsynaptic targets in either the ipsilateral pedal ganglion or the contralateral cerebropleural ganglion. Type I interneurons exhibited unique intraganglionic projections to different regions of the cerebropleural ganglion, depending on whether they received excitatory or inhibitory synaptic input from identified photoreceptors. Type I interneurons that received monosynaptic excitatory input from identified B photoreceptors terminated near the cerebropleural commissure and had multiple regions of varicosities located at branches that projected from the primary axon. Type I interneurons that received monosynaptic inhibitory input from identified B photoreceptors projected to the anterior cerebropleural ganglion and exhibited varicosities localized to the terminal region of the primary axonal process. Type II interneurons that received polysynaptic inhibitory input from identified photoreceptors projected to the contralateral cerebropleural ganglion. Most type II interneurons that projected to the pedal ganglia received polysynaptic excitatory input from identified photoreceptors. These results indicate that there is at least one additional interneuron in the higher-order visual circuit between type I interneurons and pedal motor neurons responsible for the generation of phototactic locomotion in Hermissenda.     

Influences of cortico-tectal and intertectal connections on visual responses in the cat's superior colliculus

Summary Our experiments, utilizing electrical shocks applied to the lateral- or supra-sylvian gyrus of the cortex, demonstrate an initially excitatory (latency 2–10 msec) but predominantly inhibitory influence of cortico-tectal afferents on the discharge of tectal neurons. Primary or secondary inhibition in tectal cells after cortical stimulation suppressed spontaneous or visually driven activity and limited the frequency of stimulation which tectal neurons could follow.The main influence of the contralateral colliculus on visual responses of tectal cells is inhibitory but again some principally monosynaptic intertectal connections evoked initial excitation (latency 3–10 msec) after electrical stimulation of the contralateral optic tract.Removal of the visual areas 17, 18 and 19 did not cause a loss of movement- or direction-selectivity in neurons of the superior colliculus. Cooling of the occipital cortex, while recording from direction-selective tectal neurons did not alter their essential response characteristics. The response to cortical shocks disappeared in tectal neurons during cooling but could be restored by rewarming of the cortex.It could not be confirmed in our experiments that excitation and movement- or direction-selectivity of neurons in the superior colliculus depend on a specific input from areas 17, 18 and 19 of the cortex.     

Shunting inhibition in accessory optic system neurons

The interaction of excitatory and inhibitory inputs to the accessory optic system was studied with whole cell recordings in the turtle basal optic nucleus. Previous studies have shown that visual patterns, drifting in the same preferred direction, evoke excitatory and inhibitory postsynaptic events simultaneously. Analysis of the reversal potentials for these events and their pharmacological profile suggest that they are mediated by AMPA and GABA(A) receptors, respectively. Here, neurons were recorded to study nonlinear interaction between excitatory and inhibitory responses evoked by electrical microstimulation of the retina and pretectum, respectively. The responses to coincident activation of excitatory and inhibitory inputs exhibited membrane shunting in that the excitatory response amplitude, adjusted for changes in driving force, was attenuated during the onset of the inhibitory response. This nonlinear interaction was seen in many but not all stimulus pairings. In some cases, attenuation was followed by an augmentation of the excitatory response. For comparison, the size of the excitatory response was evaluated during a hyperpolarizing current pulse that directly modulated voltage-sensitive channels of a slow rectifying I(h) current. Injection of hyperpolarizing current did not cause the attenuation of the excitatory synaptic responses. We conclude that there is a nonlinear interaction between these excitatory and inhibitory synaptic currents that is not due to hyperpolarization itself, but probably is a result of their own synaptic conductance changes, i.e., shunting. Since these events are evoked by identical visual stimuli, this interaction may play a role in visual processing.     

Synaptic activity in chronically injured,epileptogenic sensory-motor neocortex

We recorded spontaneous and evoked synaptic currents in pyramidal neurons of layer V in chronically injured, epileptogenic neocortex to assess changes in the efficacy of excitatory and inhibitory neurotransmission that might promote cortical hyperexcitability. Partial sensory-motor neocortical isolations with intact blood supply ("undercuts") were made in 20 rats on postnatal day 21-25 and examined 2-6 wk later in standard brain slice preparations using whole cell patch-clamp techniques. Age-matched, uninjured naive rats (n = 20) were used as controls. Spontaneous and miniature excitatory and inhibitory postsynaptic currents (s- and mEPSCs; s- and mIPSCs) were recorded using patch-clamp techniques. The average frequency of s- and mEPSCs was significantly higher, while that of s- and mIPSCs was significantly lower in neurons of undercuts versus controls. The increased frequency of excitatory events was due to an increase in both s- and mEPSC frequency, suggesting an increased number of excitatory contacts and/or increased release probability at excitatory terminals. No significant difference was observed in 10-90% rise time of these events. The input-output slopes of fast, short-latency, alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid/kainate (AMPA/KA) receptor-mediated components of evoked EPSCs were steeper in undercuts than in controls. The peak amplitude of the AMPA/KA component of EPSCs evoked by supra-threshold stimuli was significantly greater in the partially isolated neocortex. In contrast, the N-methyl-D-aspartate receptor-mediated component of evoked EPSCs was not significantly different in neurons of injured versus control cortex, suggesting that the increased AMPA/KA component was due to postsynaptic alterations. Results support the conclusion that layer V pyramidal neurons receive increased AMPA/KA receptor-mediated excitatory synaptic drive and decreased GABA(A) receptor-mediated inhibition in this chronically injured, epileptogenic cortex. This shift in the balance of excitatory and inhibitory synaptic activation of layer V pyramidal cells toward excitation might be maladaptive and play a critical role in epileptogenesis.     

Protracted postnatal development of inhibitory synaptic transmission in rat hippocampal area CA1 neurons

In the CNS, inhibitory synaptic function undergoes profound transformation during early postnatal development. This is due to variations in the subunit composition of subsynaptic GABA(A) receptors (GABA(A)Rs) at differing developmental stages as well as other factors. These include changes in the driving force for chloride-mediated conductances as well as the quantity and/or cleft lifetime of released neurotransmitter. The present study was undertaken to investigate the nature and time course of developmental maturation of GABAergic synaptic function in hippocampal CA1 pyramidal neurons. In neonatal [postnatal day (P) 1-7] and immature (P8-14) CA1 neurons, miniature inhibitory postsynaptic currents (mIPSCs) were significantly larger, were less frequent, and had slower kinetics compared with mIPSCs recorded in more mature neurons. Adult mIPSC kinetics were achieved by the third postnatal week in CA1 neurons. However, despite this apparent maturation of mIPSC kinetics, significant differences in modulation of mIPSCs by allosteric agonists in adolescent (P15-21) neurons were still evident. Diazepam (1-300 nM) and zolpidem (200 nM) increased the amplitude of mIPSCs in adolescent but not adult neurons. Both drugs increased mIPSC decay times equally at both ages. These differential agonist effects on mIPSC amplitude suggest that in adolescent CA1 neurons, inhibitory synapses operate differently than adult synapses and function as if subsynaptic receptors are not fully occupied by quantal release of GABA. Rapid agonist application experiments on perisomatic patches pulled from adolescent neurons provided additional support for this hypothesis. In GABA(A)R currents recorded in these patches, benzodiazepine amplitude augmentation effects were evident only when nonsaturating GABA concentrations were applied. Furthermore nonstationary noise analysis of mIPSCs in P15-21 neurons revealed that zolpidem-induced mIPSC augmentation was not due to an increase in single-channel conductance of subsynaptic GABA(A)Rs but rather to an increase in the number of open channels responding to a single GABA quantum, further supporting the hypothesis that synaptic receptors may not be saturated during synaptic function in adolescent neurons. These data demonstrate that inhibitory synaptic transmission undergoes a markedly protracted postnatal maturation in rat CA1 pyramidal neurons. In the first two postnatal weeks, mIPSCs are large in amplitude, are slow, and occur infrequently. By the third postnatal week, mIPSCs have matured kinetically but retain distinct responses to modulatory drugs, possibly reflecting continued immaturity in synaptic structure and function persisting through adolescence.     

Transient enhancement of inhibitory synaptic transmission in hippocampal CA1 pyramidal neurons after cerebral ischemia

Pyramidal neurons in hippocampal CA1 regions are highly sensitive to cerebral ischemia. Alterations of excitatory and inhibitory synaptic transmission may contribute to the ischemia-induced neuronal degeneration. However, little is known about the changes of GABAergic synaptic transmission in the hippocampus following reperfusion. We examined the GABA_A receptor-mediated inhibitory postsynaptic currents (IPSCs) in CA1 pyramidal neurons 12 and 24 h after transient forebrain ischemia in rats. The amplitudes of evoked inhibitory postsynaptic currents (eIPSCs) were increased significantly 12 h after ischemia and returned to control levels 24 h following reperfusion. The potentiation of eIPSCs was accompanied by an increase of miniature inhibitory postsynaptic current (mIPSC) amplitude, and an enhanced response to exogenous application of GABA, indicating the involvement of postsynaptic mechanisms. Furthermore, there was no obvious change of the paired-pulse ratio (PPR) of eIPSCs and the frequency of mIPSCs, suggesting that the potentiation of eIPSCs might not be due to the increased presynaptic release. Blockade of adenosine A1 receptors led to a decrease of eIPSCs amplitude in post-ischemic neurons but not in control neurons, without affecting the frequency of mIPSCs and the PPR of eIPSCs. Thus, tonic activation of adenosine A1 receptors might, at least in part, contribute to the enhancement of inhibitory synaptic transmission in CA1 neurons after forebrain ischemia. The transient enhancement of inhibitory neurotransmission might temporarily protect CA1 pyramidal neurons, and delay the process of neuronal death after cerebral ischemia.     

Magnocellular and parvocellular divisions of pigeon nucleus isthmi differentially modulate visual responses in the tectum

The electrophysiological responses of 162 tectal cells to computer-generated visual stimuli were extracellularly recorded from 24 homing pigeons before and after injecting either lidocaine or N-methyl-d-aspartate (NMDA) into the nucleus isthmi pars magnocellularis (Imc) or the nucleus isthmi pars parvocellularis (Ipc). Micro-injections of lidocaine into Imc resulted in a significant reduction of firing rate in 80% of tectal cells, whose excitatory receptive fields (ERFs) were localized within the ERF of the Imc cell where the lidocaine was injected. In contrast, when lidocaine was injected into Ipc under identical circumstances it had no effect on the visually driven activity of 68% of tectal cells. However, when the excitatory amino acid NMDA was injected into Ipc it produced a significant reduction in the visually driven firing of 75% of tectal neurons when their ERFs were within the isthmic ERF, while similar application of NMDA into Imc had no effect on the visually driven response of 94% of tectal neurons. When the ERFs of tectal cells were localized outside the ERF of the isthmic cell where the chemical was injected, Imc-injected lidocaine had no effect in 9 out of 10 tectal cells, whereas Ipc-injected NMDA increased firing in 7 out of 17 tectal cells. Therefore, it is suggested that the Imc-tectal fibers participate in a positive feedback pathway and the Ipc-tectal fibers are involved in a negative feedback pathway.     

Long-term actions of BDNF on inhibitory synaptic transmission in identified neurons of the rat substantia gelatinosa

Peripheral nerve injury promotes the release of brain-derived neurotrophic factor (BDNF) from spinal microglial cells and primary afferent terminals. This induces an increase in dorsal horn excitability that contributes to "central sensitization" and to the onset of neuropathic pain. Although it is accepted that impairment of GABAergic and/or glycinergic inhibition contributes to this process, certain lines of evidence suggest that GABA release in the dorsal horn may increase after nerve injury. To resolve these contradictory findings, we exposed rat spinal cord neurons in defined-medium organotypic culture to 200 ng/ml BDNF for 6 days to mimic the change in spinal BDNF levels that accompanies peripheral nerve injury. Morphological and electrophysiological criteria and glutamic acid decarboxylase (GAD) immunohistochemistry were used to distinguish putative inhibitory tonic-islet-central neurons from putative excitatory delay-radial neurons. Whole cell recording in the presence of 1 μM tetrodotoxin showed that BDNF increased the amplitude of GABAergic and glycinergic miniature inhibitory postsynaptic currents (mIPSCs) in both cell types. It also increased the amplitude and frequency of spontaneous, action potential-dependent IPSCs (sIPSCs) in putative excitatory neurons. By contrast, BDNF reduced sIPSC amplitude in inhibitory neurons but frequency was unchanged. This increase in inhibitory drive to excitatory neurons and decreased inhibitory drive to inhibitory neurons seems inconsistent with the observation that BDNF increases overall dorsal horn excitability. One of several explanations for this discrepancy is that the action of BDNF in the substantia gelatinosa is dominated by previously documented increases in excitatory synaptic transmission rather than by impediment of inhibitory transmission.     

Inhibitory synaptic plasticity regulates pyramidal neuron spiking in the rodent hippocampus

Spike-timing modifies the efficacy of both excitatory and inhibitory synapses onto CA1 pyramidal neurons in the rodent hippocampus. Repetitively spiking the presynaptic neuron before the postsynaptic neuron induces inhibitory synaptic plasticity, which results in a depolarization of the reversal potential for GABA (E(GABA)). Our goal was to determine how inhibitory synaptic plasticity regulates CA1 pyramidal neuron spiking in the rat hippocampus. We demonstrate electrophysiologically that depolarizing E(GABA) by 24.7 mV increased the spontaneous action potential firing frequency of cultured hippocampal neurons 254% from 0.12+/-0.07 Hz to 0.44+/-0.13 Hz (n=11; P<0.05). Next we used a single compartment model of a CA1 pyramidal neuron to explore in detail how inhibitory synaptic plasticity of feedforward and feedback inhibition regulates the generation of action potentials, spike latency, and the minimum excitatory conductance required to generate an action potential; plasticity was modeled as a depolarization of E(GABA), which effectively weakens inhibition. Depolarization of E(GABA) at feedforward and feedback inhibitory synapses decreased the latency to the 1st spike by 2.27 ms, which was greater that the sum of the decreases produced by depolarizing E(GABA) at feedforward (0.85 ms) or feedback inhibitory synapses (0.02 ms) alone. In response to a train of synaptic inputs, depolarizing E(GABA) decreased the inter-spike interval and increased the number of output spikes in a frequency dependent manner, improving the reliability of input-output transmission. Moreover, a depolarizing shift in E(GABA) at feedforward and feedback synapses triggered by spike trains recorded from CA1 pyramidal layer neurons during field theta from anesthetized rats, significantly increased spiking on the up- and down-strokes of the first half of the theta rhythm (P<0.05), without changing the preferred phase of firing (P=0.783). This study provides the first explanation of how depolarizing E(GABA) affects pyramidal cell output within the hippocampus.     

Dynamic response properties of visual neurons and context-dependent surround effects on receptive fields in the tectum of the salamander Plethodon shermani

Neuronal responses to complex prey-like stimuli and rectangles were investigated in the tectum of the salamander Plethodon shermani using extracellular single-cell recording. Cricket dummies differing in size, contrast or movement pattern or a rectangle were moved singly through the excitatory receptive field of a neuron. Paired presentations were performed, in which a reference stimulus was moved inside and the different cricket dummies or the rectangle outside the excitatory receptive field. Visual object recognition involves much more complex spatial and temporal processing than previously assumed in amphibians. This concerns significant changes in absolute number of spikes, temporal discharge pattern, and receptive field size. At single presentation of stimuli, the number of discharges was significantly changed compared with the reference stimulus, and in the majority of neurons the temporal pattern of discharges was changed in addition. At paired presentation of stimuli, neurons mainly revealed a significant decrease in average spike number and a reduction of excitatory receptive field size to presentation of the reference stimulus inside the excitatory receptive field, when a large-sized cricket stimulus or the rectangle was located outside the excitatory receptive field. This inhibition was significantly greater for the large-sized cricket stimulus than for the rectangle, and indicates the biological relevance of the prey-like stimulus in object selection. The response properties of tectal neurons at single or paired presentation of stimuli indicate that tectal neurons integrate information across a much larger part of visual space than covered by the excitatory receptive field. The spike number of a tectal neuron and the spatio-temporal extent of its excitatory receptive field are not fixed but depend on the context, i.e. the stimulus type and combination. This dynamic processing corresponds with the selection of the stimuli in the visual orienting behavior of Plethodon investigated in a previous study, and we assume that tectal processing is modulated by top down processes as well as feedback circuitries.     

Spontaneous synaptic activity is primarily GABAergic in vestibular nucleus neurons of the chick embryo

The principal cells of the chick tangential nucleus are vestibular nucleus neurons participating in the vestibular reflexes. In 16-day embryos, the application of glutamate receptor antagonists abolished the postsynaptic responses generated on vestibular-nerve stimulation, but spontaneous synaptic activity was largely unaffected. Here, spontaneous synaptic activity was characterized in principal cells from brain slices at E16 using whole cell voltage-clamp recordings. With KCl electrodes, the frequency of spontaneous inward currents was 3.1 Hz at -60 mV, and the reversal potential was +4 mV. Cs-gluconate pipette solution allowed the discrimination of glycine/GABA(A) versus glutamate receptor-mediated events according to their different reversal potentials. The ratio for spontaneous excitatory to inhibitory events was about 1:4. Seventy-four percent of the outward events were GABA(A), whereas 26% were glycine receptor-mediated events. Both pre- and postsynaptic GABA(B) receptor effects were shown, with presynaptic GABA(B) receptors inhibiting 40% of spontaneous excitatory postsynaptic currents (sEPSCs) and 53% of spontaneous inhibitory postsynaptic currents (sIPSCs). With TTX, the frequency decreased approximately 50% for EPSCs and 23% for IPSCs. These data indicate that the spontaneous synaptic activity recorded in the principal cells at E16 is primarily inhibitory, action potential-independent, and based on the activation of GABA(A) receptors that can be modulated by presynaptic GABA(B) receptors.     

Effects of glutamatergic, cholinergic and gabaergic antagonists on tectal cells in toads

The present paper using microiontophoresis analysis describes transmitters and their receptor subtypes used in retinotectal and isthmotectal transmission, and suggests several modes converging retinotectal and isthmotectal afferents on tectal neurons in toads (Bufo bufo gargarizans). Neuronal responses of tectal cells were extracellularly recorded to both visual stimulation and electrical stimulation of the nucleus isthmi, and effects of glutamatergic, cholinergic, GABAergic and glycinergic antagonists on these responses examined. Visual responses in 80% of tectal cells were reversibly blocked by the N-methyl-D-aspartate antagonist 3-Rs-2-carboxypiperazin-4-yl-propyl-1-phosphonic acid, and those of the remaining 20% of cells by the muscarinic antagonist atropine, suggesting that there may be at least two kinds of retinotectal synapse that use glutamate and N-methyl-D-aspartate receptors, and acetylcholine and muscarinic receptors, respectively. Electrical stimulation of the nucleus isthmi elicited excitatory responses in 67% of tectal cells, excitatory-inhibitory responses in 16% of cells, and inhibitory responses in 17% of cells examined. The excitatory responses were reversibly abolished by atropine, but not affected by either 3-Rs-2-carboxypiperazin-4-yl-propyl-1-phosphonic acid or the alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate antagonist 6-cyano-7-nitroquinoxaline-2,3-dione, whereas the inhibitory responses were released by the GABA receptor A antagonist bicuculline, but not influenced by the GABA receptor B antagonist 2-hydroxysaclofen and glycinergic antagonist strychnine. Excitatory and inhibitory components in the excitatory-inhibitory responses were blocked by atropine and bicuculline, respectively. It appears that glutamatergic and cholinergic afferents from the retina, and cholinergic and GABAergic afferents from the nucleus isthmi may converge on tectal neurons in at least five modes of synaptic connections, in agreement with the heterogeneous populations of tectal cells in amphibians.     

Direction tuning of inhibitory inputs to the turtle accessory optic system.

Neurons in turtle accessory optic system (basal optic nucleus, BON) were studied to compare excitatory and inhibitory visual inputs. Using a reduced in vitro brain stem preparation with the eyes attached, previous studies only showed a monosynaptic retinal input to the BON from direction-sensitive retinal ganglion cells that share a common preferred direction. Now using an intact brain stem preparation, not only did BON neurons display inhibitory postsynaptic potentials [IPSP(C)s] spontaneously, but IPSP(C)s were also evoked by visual pattern motion, they had their polarity reversed near the chloride equilibrium potential and they were blocked by the GABA(A) antagonist bicuculline. Because excitatory postsynaptic currents had reversal potentials >0 mV, BON cells were recorded using patch electrodes filled with QX-314 or Cs+ to measure the cell's direction tuning also at that higher reversal potential. For most of the BON neurons studied, their visual excitation and inhibition had a very similar preferred direction, indicating that both synaptic inputs were maximally active onto the same cell under the same stimulus conditions. These competing inputs may result from connections between the pretectum and accessory optic nuclei. Such synaptic interactions may serve a functional role in the visual processing necessary to create retinal slip signals for oculomotor control.     

Inhibitory synaptic transmission differs in mouse type A and B medial vestibular nucleus neurons in vitro

Fast inhibitory synaptic transmission in the medial vestibular nucleus (MVN) is mediated by GABA(A) receptors (GABA(A)Rs) and glycine receptors (GlyRs). To assess their relative contribution to inhibition in the MVN, we recorded miniature inhibitory postsynaptic currents (mIPSCs) in physiologically characterized type A and type B MVN neurons. Transverse brain stem slices were prepared from mice (3-8 wk old), and whole cell patch-clamp recordings were obtained from visualized MVN neurons (CsCl internal; Vm = -70 mV; 23 degrees C). In 81 MVN neurons, 69% received exclusively GABA(A)ergic inputs, 6% exclusively glycinergic inputs, and 25% received both types of mIPSCs. The mean amplitude of GABA(A)R-mediated mIPSCs was smaller than those mediated by GlyRs (22.6 +/- 1.8 vs. 35.3 +/- 5.3 pA). The rise time and decay time constants of GABA(A)R- versus GlyR-mediated mIPSCs were slower (1.3 +/- 0.1 vs. 0.9 +/- 0.1 ms and 10.5 +/- 0.3 vs. 4.7 +/- 0.3 ms, respectively). Comparison of type A (n = 20) and type B (n = 32) neurons showed that type A neurons received almost exclusively GABA(A)ergic inhibitory inputs, whereas type B neurons received GABA(A)ergic inputs, glycinergic inputs, or both. Intracellular labeling in a subset of MVN neurons showed that morphology was not related to a MVN neuron's inhibitory profile (n = 15), or whether it was classified as type A or B (n = 29). Together, these findings indicate that both GABA and glycine contribute to inhibitory synaptic processing in MVN neurons, although GABA dominates and there is a difference in the distribution of GABA(A) and Gly receptors between type A and type B MVN neurons.     

Transition from GABAergic to glycinergic synaptic transmission in newly formed spinal networks.

The role of glycinergic and GABAergic systems in mediating spontaneous synaptic transmission in newly formed neural networks was examined in motoneurons in the developing rat spinal cord. Properties of action potential-independent miniature inhibitory postsynaptic currents (mIPSCs) mediated by glycine and GABA(A) receptors (GlyR and GABA(A)R) were studied in spinal cord slices of 17- to 18-day-old embryos (E17-18) and 1- to 3-day-old postnatal rats (P1-3). mIPSC frequency and amplitude significantly increased after birth, while their decay time decreased. To determine the contribution of glycinergic and GABAergic synapses to those changes, GlyR- and GABA(A)R-mediated mIPSCs were isolated based on their pharmacological properties. Two populations of pharmacologically distinct mIPSCs were recorded in the presence of glycine or GABA(A) receptors antagonists: bicuculline-resistant, fast-decaying GlyR-mediated mIPSCs, and strychnine-resistant, slow-decaying GABA(A)R-mediated mIPSCs. The frequency of GABA(A)R-mediated mIPSCs was fourfold higher than that of GlyR-mediated mIPSCs at E17-18, indicating that GABAergic synaptic sites were functionally dominant at early stages of neural network formation. Properties of GABA(A)R-mediated mIPSC amplitude fluctuations changed from primarily unimodal skewed distribution at E17-18 to Gaussian mixtures with two to three discrete components at P1-3. A developmental shift from primarily long-duration GABAergic mIPSCs to short-duration glycinergic mIPSCs was evident after birth, when the frequency of GlyR-mediated mIPSCs increased 10-fold. This finding suggested that either the number of glycinergic synapses or the probability of vesicular glycine release increased during the period studied. The increased frequency of GlyR-mediated mIPSCs was associated with more than a twofold increase in their mean amplitude, and in the number of motoneurons in which mIPSC amplitude fluctuations were best fitted by multi-component Gaussian curves. A third subpopulation of mIPSCs was apparent in the absence of glycine and GABA(A) receptor antagonists: mIPSCs with both fast and slow decaying components. Based on their dual-component decay time and their suppression by either strychnine or bicuculline, we assumed that these were generated by the activation of co-localized postsynaptic glycine and GABA(A) receptors. The contribution of mixed glycine-GABA synaptic sites to the generation of mIPSCs did not change after birth. The developmental switch from predominantly long-duration GABAergic inhibitory synaptic currents to short-duration glycinergic currents might serve as a mechanism regulating neuronal excitation in the developing spinal networks.     

Distribution of GABA immunoreactivity in the optic tectum of the frog: a light and electron microscopic study

GABA immunoreactivity was studied in the optic tectum of the frog, Rana esculenta, by postembedding immunohistochemical methods at the light and electron microscopic levels. Nearly one-third of the total population of tectal cells appeared to be GABA-immunoreactive. The proportion of stained neurons was highest in layer 9 (61%), and they occurred less frequently in layers 7 (21%) and 6 (27%). Stained perikarya represented a population of small neurons with a diameter of 8-10 microns. Large cell bodies in layer 7 or at the top of layer 6, and cells of origin of the mesencephalic trigeminal tract in layer 2, were devoid of labelling. Axon terminals and dendrites displaying immunoreactivity for GABA were observed in all of the plexiform layers. On the basis of ultrastructural characteristics two types of GABA-positive axon terminals and two variations of GABA-immunoreactive dendrites were distinguished. Synaptic relations of GABA-immunoreactive and GABA-negative axons as well as dendrites were also studied. Besides a wide variety of axodendritic synapses, dendrodendritic synaptic appositions were also revealed. The results suggest that various inhibitory mechanisms are involved in tectal circuits, which have to be incorporated into future neuronal models concerning visual information processing in the optic tectum of the frog.     

Spontaneous synaptic activity in chick vestibular nucleus neurons during the perinatal period

The principal cells of the chick tangential nucleus are second-order vestibular neurons involved in the vestibuloocular and vestibulocollic reflexes. The spontaneous synaptic activity of morphologically identified principal cells was characterized in brain slices from 1-day-old hatchlings (H1) using whole-cell voltage-clamp recordings and Cs-gluconate pipet solution. The frequency was 1.45 Hz for spontaneous excitatory postsynaptic currents (sEPSCs) and 1.47 Hz for spontaneous inhibitory postsynaptic currents (sIPSCs). Using specific neurotransmitter receptor antagonists, all of the sEPSCs were identified as AMPA receptor-mediated events, whereas 56% of the sIPSCs were glycine and 44% were GABA(A) receptor-mediated events. On exposure to TTX, the frequency of EPSCs decreased by 68%, while the frequency of IPSCs decreased by 33%, indicating greater EPSC dependency on presynaptic action potentials. These data on spontaneous synaptic activity at H1 were compared with those obtained in previous studies of 16-day old embryos (E16). After birth, the spontaneous synaptic activity exhibited increased EPSC frequency, increased ratio for excitatory to inhibitory events, increased percentage of TTX-dependent EPSCs, and faster kinetics. In addition, the ratio for glycine/GABA receptor-mediated events increased significantly. Altogether, these data indicate that at hatching spontaneous synaptic activity of vestibular nucleus neurons in brain slices of the chick tangential nucleus undergoes appreciable changes, with increased frequency of EPSCs and glycinergic activity playing more important roles compared with the late-term chick embryo when GABAergic activity prevailed. The definition of this developmental pattern of synaptic activity in vestibular nucleus neurons should contribute to understanding how vestibular reflex activity is established in the hatchling chick.     

Differences between the scaling of miniature IPSCs and EPSCs recorded in the dendrites of CA1 mouse pyramidal neurons

Anatomical studies have described inhibitory synaptic contacts on apical dendrites, and an abundant number of GABAergic synapses on the somata and proximal dendrites of CA1 pyramidal cells of the hippocampus. The number of inhibitory contacts decreases dramatically with distance from the soma, but the local electrophysiological characterization of these synapses at their site of origin in the dendrites is missing. We directly recorded dendritic GABA receptor-mediated inhibitory synaptic events in adult mouse hippocampal CA1 pyramidal neurons and compared them to excitatory synaptic currents recorded at the same sites. Miniature GABAergic events were evoked using localized application of a hyperosmotic solution to the apical dendrites in the vicinity of the dendritic whole-cell recording pipette. Glutamatergic synaptic events were blocked by kynurenic acid, leaving picrotoxin-sensitive IPSCs. We measured the amplitude and kinetic properties of mIPSCs at the soma and at three different dendritic locations. The amplitude of mIPSCs recorded at the various sites was similar along the somato-dendritic axis. The rise- and decay-times of local mIPSCs were also independent of the location of the synapses. The frequency of mIPSCs was 5 Hz at the soma, in contrast to < 0.5 Hz at dendritic sites, which could be increased to 10–20 Hz and 6–10 Hz, respectively, by our hyperosmotic stimulation protocol. Miniature glutamatergic events were evoked with the same protocol after blocking inhibitory synapses by bicucculine. The measured amplitudes increased along the somato-dendritic axis proportionally with their distance from the soma. The measured kinetic properties were independent of location. Consistent with the idea that IPSCs may have a restricted local effect in the dendrites, our data show a lack of distance-dependent scaling of miniature inhibitory synaptic events, in contrast to the scaling of excitatory events recorded at the same sites.     

A role of insulin-like growth factor 1 in beta amyloid-induced disinhibition of hippocampal neurons

In the present study we investigated the effects of beta amyloid (Abeta) on inhibitory synaptic transmission in the cultured hippocampal neurons using whole-cell patch-clamp recordings and immunocytochemistry, and examined the role of insulin-like growth factor 1 (IGF-1). Incubation with 4 microM Abeta25-35 for 24 h significantly decreased the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs), but had no effect on the mean amplitude. Pretreatment with 10 ng/ml IGF-1 for 24h prior to Abeta25-35 exposure blocked Abeta-induced disinhibition of hippocampal neurons. The frequency and mean amplitude of miniature IPSC (mIPSCs) were not significantly affected by Abeta. The rise and decay kinetics of sIPSCs and mIPSCs were similar for the control and Abeta25-35-treated hippocampal neurons. Immunocytochemistry showed no changes in the ratio of gamma-aminobutyric acid (GABA) positive cells subsequent to treatment with Abeta, or IGF-1. Together these data suggest that Abeta-induced the disinhibition in cultured hippocampal neurons, whereas IGF-1 could block this effect.