Intracellular blockade of GABAA receptors in the rat hippocampal neurons

The intracellular blockade of GABA_A-receptor-mediated currents is a useful approach to suppress the GABAergic conductance in a single cell and to isolate the glutamatergic component of network-driven activities. Previously an approach has been described allowing intracellular blockade of GABA_A receptors by means of intracellular dialysis of a neuron with the pipette-filling solution, in which fluoride ions that hardly pass through the GABA_A receptor channels substitute for Cl^? and in which Mg²⁺ and ATP are omitted to induce rundown of the GABA_A receptors during whole-cell patch-clamp recordings. However, the kinetics of suppression of GABAergic conductance and the effect on the currents mediated by glutamate receptors remain unknown. Here, using whole-cell recordings with fluoride-based, Mg²⁺- and ATP-free solution on CA3 hippocampal neurons of neonatal rats, we show that after 1 h of such dialysis, both spontaneous and evoked GABA_A-receptor-mediated synaptic currents and responses induced by the GABA_A receptor agonist isoguvacine were completely suppressed. Inward GABAergic postsynaptic currents were suppressed prior to outward currents. Synaptic responses mediated by AMPA receptors were not affected by the dialysis, whereas the NMDA-receptor-mediated postsynaptic currents were reduced by approximately 20%. Dialysis with fluoride-based Mg²⁺, ATP-free solution either fully blocked giant depolarizing potentials (GDPs) in CA3 pyramidal cells (n = 2) or reduced the charge crossing the membrane during GDPs and shifted the GDP reversal potential to more positive values (n = 5). The dialysis-resistant component of GDPs was mediated by glutamate receptors, since: (i) it reversed around 0 mV; (ii) it demonstrated a negative slope conductance at negative membrane voltages, which is characteristic of NMDA receptor-mediated responses; (iii) kinetics of the individual events composing the dialysis-resistant component of GDPs at negative voltages were very similar to those of AMPA receptor-mediated synaptic currents. Thus, this procedure can be useful to isolate the glutamate receptor-mediated component of neuronal network-driven activities.     

Excitatory actions of GABA in the cortex

Little is known about how GABAergic inputs interact with excitatory inputs under conditions that maintain physiological concentrations of intracellular anions. Using extracellular and gramicidin perforated-patch recording, we show that somatic and dendritic GABA responses in mature cortical pyramidal neurons are depolarizing from rest and can facilitate action potential generation when combined with proximal excitatory input. Dendritic GABA responses were excitatory regardless of timing, whereas somatic GABA responses were inhibitory when coincident with excitatory input but excitatory at earlier times. These excitatory actions of GABA occur even though the GABA reversal potential is below action potential threshold and largely uniform across the somato-dendritic axis, and arise when GABAergic inputs are temporally or spatially isolated from concurrent excitation. Our findings demonstrate that under certain circumstances GABA will have an excitatory role in synaptic integration in the cortex.     

Neurochemistry of GABAergic activities in the central nervous system ofLocusta migratoria

Summary The biochemical elements of GABA-ergic synapses in the central nervous tissue were examined by a comparative neurochemical approach. The high concentration of GABA as well as the activities of glutamate decarboxylase and GABA-transaminase suppose a high content of GABAergic elements in the nervous system of the locust.Nerve endings isolated from the ganglia of locusts accumulated exogenous GABA in a carriermediated, sodium dependent process into compartments from where it could partially be released under depolarizing conditions. The transport was stimulated by extracellular chloride, was modulated by specific ionophores (enhanced by valinomycin, inhibited by CCCP) and could effectively be blocked by GABAergic ligands (DABA, muscimol). Binding studies revealed the existence of multiple binding sites for GABA which differ in number, affinity, pharmacology and ion dependency. The putative receptors for GABA (Na⁺-independent binding sites) in locust nervous tissue exceeded the concentrations found in vertebrate brain tissue and showed different binding pharmacology.Abbreviations GABA -amino butyric acid - GAD glutamate decarboxylase - GABA-T GABA-transaminase - DABA diamino butyric acid     

Postnatal maturation of gamma-aminobutyric acidA and B-mediated inhibition in the CA3 hippocampal region of the rat

In the adult central nervous system, GABAergic synaptic inhibition is known to play a crucial role in preventing the spread of excitatory glutamatergic activity. This inhibition is achieved by a membrane hyperpolarization through the activation of postsynaptic γ-aminobutyric acid_A (GABA_A) and GABA_B receptors. In addition, GABA also depress transmitter release acting through presynaptic GABA_B receptors. Despite the wealth of data regarding the role of GABA in regulating the degree of synchronous activity in the adult, little is known about GABA transmission during early stages of development. In the following we report that GABA mediates most of the excitatory drive at early stages of development in the hippocampal CA3 region. Activation of GABA_A receptors induces a depolarization and excitation of immature CA3 pyramidal neurons and increases intracellular Ca²⁺ ([Ca²⁺]_i) during the first postnatal week of life. During the same developmental period, the postsynaptic GABA_B-mediated inhibition is poorly developed. In contrast, the presynaptic GABA_B-mediated inhibition is well developed at birth and plays a crucial role in modulating the postsynaptic activity by depressing transmitter release at early postnatal stages. We have also shown that GABA plays a trophic role in the neuritic outgrowth of cultured hippocampal neurons. © 1995 John Wiley & Sons, Inc.     

The synaptic scaffold protein MPP2 interacts with GABAA receptors at the periphery of the postsynaptic density of glutamatergic synapses

Recent advances in imaging technology have highlighted that scaffold proteins and receptors are arranged in subsynaptic nanodomains. The synaptic membrane-associated guanylate kinase (MAGUK) scaffold protein membrane protein palmitoylated 2 (MPP2) is a component of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor–associated protein complexes and also binds to the synaptic cell adhesion molecule SynCAM 1. Using superresolution imaging, we show that—like SynCAM 1—MPP2 is situated at the periphery of the postsynaptic density (PSD). In order to explore MPP2-associated protein complexes, we used a quantitative comparative proteomics approach and identified multiple γ-aminobutyric acid (GABA)_A receptor subunits among novel synaptic MPP2 interactors. In line with a scaffold function for MPP2 in the assembly and/or modulation of intact GABA_A receptors, manipulating MPP2 expression had effects on inhibitory synaptic transmission. We further show that GABA_A receptors are found together with MPP2 in a subset of dendritic spines and thus highlight MPP2 as a scaffold that serves as an adaptor molecule, linking peripheral synaptic elements critical for inhibitory regulation to central structures at the PSD of glutamatergic synapses.

This study shows that the MAGUK scaffold protein MPP2 is located at the periphery of postsynaptic densities in excitatory neurons, where it interacts with GABA-A receptors, thereby serving as a functional adaptor that links excitatory and inhibitory components of synaptic transmission at glutamatergic synapses.     

Phase Dependent Sign Changes of GABAergic Synaptic Input Explored <Emphasis Type="Italic">In-Silicio</Emphasis> and <Emphasis Type="Italic">In-Vitro</Emphasis>

Inhibitory interactions play a crucial role in the synchronization of neuronal activity. Here we investigate the effect of GABAergic PSPs on spike timing in cortical neurons that exhibit an oscillatory modulation of their membrane potential. To this end we combined numerical simulations with in-vitro patch-clamp recordings from layer II/III pyramidal cells of the rat visual cortex. Special emphasis was placed on exploring how the reversal potential of the GABAergic synaptic currents (E_GABA) and the phase relations of the PSPs relative to the oscillation cycles affect the timing of spikes riding on the depolarizing peaks of the oscillations. The simulations predicted: (1) With E_GABA more negative than the oscillation minima PSPs are hyperpolarizing at all phases and thus delay or prevent spikes. (2) With E_GABA being more positive than the oscillation maxima PSPs are depolarizing in a phase-independent way and lead to a phase advance of spikes. (3) In the intermediate case where E_GABA lies within oscillation maxima and minima PSPs are either hyper- or depolarizing depending on their phase relations to the V_m oscillations and can therefore either delay or advance spikes. Experiments conducted in this most interesting last configuration with biphasic PSPs agreed with the model predictions. Additional theoretical investigations revealed the effect of these PSP induced shifts in spike timing on synchronization in neuronal circuits. The results suggest that GABAergic mechanisms can assume highly specific timing functions in oscillatory networks.Action Editor: Alain Destexhe     

Calcium requirement of long-term depression and rebound potentiation in cerebellar Purkinje neurons

Cerebellar Purkinje neurons (PNs) receive two main excitatory inputs, from climbing fibers and parallel fibers, and inhibitory inputs, from GABAergic interneurons. The synapses formed by parallel fibers and by inhibitory interneurons on PNs are able to undergo long-lasting in efficacy. Thus, the excitatory parallel fiber-PN synapse undergoes long-term fibers. Synaptic inhibition can be potentiated by climbing fiber activity by a mechanism named rebound potentiation, resulting in a more powerful inhibitory effect of GABAergic interneurons. The induction of both long-term depression and rebound potentiation requires a transient elevation of the cytoplasmic calcium concentration ([Ca²⁺]_i). The [Ca²⁺]_i-transient is caused by Ca²⁺ entry through voltage-gated Ca²⁺ channels and, possibly, by release of Ca²⁺ from IP_3- and ryanodine-sensitive stores. Direct Ca²⁺ entry through synaptic AMPA receptor channels seems not to contribute significantly to the Ca²⁺ signal mediating the induction of both long-term depression and rebound potentiation.     

Computational Modeling Reveals Dendritic Origins of GABAA-Mediated Excitation in CA1 Pyramidal Neurons

GABA is the key inhibitory neurotransmitter in the adult central nervous system, but in some circumstances can lead to a paradoxical excitation that has been causally implicated in diverse pathologies from endocrine stress responses to diseases of excitability including neuropathic pain and temporal lobe epilepsy. We undertook a computational modeling approach to determine plausible ionic mechanisms of GABA_A-dependent excitation in isolated post-synaptic CA1 hippocampal neurons because it may constitute a trigger for pathological synchronous epileptiform discharge. In particular, the interplay intracellular chloride accumulation via the GABA_A receptor and extracellular potassium accumulation via the K/Cl co-transporter KCC2 in promoting GABA_A-mediated excitation is complex. Experimentally it is difficult to determine the ionic mechanisms of depolarizing current since potassium transients are challenging to isolate pharmacologically and much GABA signaling occurs in small, difficult to measure, dendritic compartments. To address this problem and determine plausible ionic mechanisms of GABA_A-mediated excitation, we built a detailed biophysically realistic model of the CA1 pyramidal neuron that includes processes critical for ion homeostasis. Our results suggest that in dendritic compartments, but not in the somatic compartments, chloride buildup is sufficient to cause dramatic depolarization of the GABA_A reversal potential and dominating bicarbonate currents that provide a substantial current source to drive whole-cell depolarization. The model simulations predict that extracellular K⁺ transients can augment GABA_A-mediated excitation, but not cause it. Our model also suggests the potential for GABA_A-mediated excitation to promote network synchrony depending on interneuron synapse location - excitatory positive-feedback can occur when interneurons synapse onto distal dendritic compartments, while interneurons projecting to the perisomatic region will cause inhibition.     

The extracellular matrix and synapses

Extracellular matrix (ECM) molecules, derived from both neurons and glial cells, are secreted and accumulate in the extracellular space to regulate various aspects of pre- and postsynaptic differentiation, the maturation of synapses, and their plasticity. The emerging mechanisms comprise interactions of agrin, integrin ligands, and reelin, with their cognate cell-surface receptors being coupled to tyrosine kinase activities. These may induce the clustering of postsynaptic receptors and changes in their composition and function. Furthermore, direct interactions of laminins, neuronal pentraxins, and tenascin-R with voltage-gated Ca²⁺ channels, α-amino-3-hydroxy-5-methylisoxazole-4-proprionic acid (AMPA), and γ-aminobutyric acid_B (GABA_B) receptors, respectively, shape the organization and function of different subsets of synapses. Some of these mechanisms significantly contribute to the induction of long-term potentiation in excitatory synapses, either by the regulation of Ca²⁺ entry via N-methyl-D-aspartate receptors or L-type Ca²⁺ channels, or by the control of GABAergic inhibition.A.D. was supported by DFG grants Di 702/4-1,-2 and -3.     

Structure of Excitatory Synapses and GABAA Receptor Localization at Inhibitory Synapses Are Regulated by Neuroplastin-65

Formation, maintenance, and activity of excitatory and inhibitory synapses are essential for neuronal network function. Cell adhesion molecules (CAMs) are crucially involved in these processes. The CAM neuroplastin-65 (Np65) highly expressed during periods of synapse formation and stabilization is present at the pre- and postsynaptic membranes. Np65 can translocate into synapses in response to electrical stimulation and it interacts with subtypes of GABA_A receptors in inhibitory synapses. Here, we report that in the murine hippocampus and in hippocampal primary culture, neurons of the CA1 region and the dentate gyrus (DG) express high Np65 levels, whereas expression in CA3 neurons is lower. In neuroplastin-deficient (Np^−/−) mice the number of excitatory synapses in CA1 and DG, but not CA3 regions is reduced. Notably this picture is mirrored in mature Np^−/− hippocampal cultures or in mature CA1 and DG wild-type (Np^+/+) neurons treated with a function-blocking recombinant Np65-Fc extracellular fragment. Although the number of GABAergic synapses was unchanged in Np^−/− neurons or in mature Np65-Fc-treated Np^+/+ neurons, the ratio of excitatory to inhibitory synapses was significantly lower in Np^−/− cultures. Furthermore, GABA_A receptor composition was altered at inhibitory synapses in Np^−/− neurons as the α1 to α2 GABA_A receptor subunit ratio was increased. Changes of excitatory and inhibitory synaptic function in Np^−/− neurons were confirmed evaluating the presynaptic release function and using patch clamp recording. These data demonstrate that Np65 is an important regulator of the number and function of synapses in the hippocampus.     

Quantitative Organization of GABAergic Synapses in the Molecular Layer of the Mouse Cerebellar Cortex

In the cerebellar cortex, interneurons of the molecular layer (stellate and basket cells) provide GABAergic input to Purkinje cells, as well as to each other and possibly to other interneurons. GABAergic inhibition in the molecular layer has mainly been investigated at the interneuron to Purkinje cell synapse. In this study, we used complementary subtractive strategies to quantitatively assess the ratio of GABAergic synapses on Purkinje cell dendrites versus those on interneurons. We generated a mouse model in which the GABA_A receptor α1 subunit (GABA_ARα1) was selectively removed from Purkinje cells using the Cre/loxP system. Deletion of the α1 subunit resulted in a complete loss of GABA_AR aggregates from Purkinje cells, allowing us to determine the density of GABA_AR clusters in interneurons. In a complementary approach, we determined the density of GABA synapses impinging on Purkinje cells using α-dystroglycan as a specific marker of inhibitory postsynaptic sites. Combining these inverse approaches, we found that synapses received by interneurons represent approximately 40% of all GABAergic synapses in the molecular layer. Notably, this proportion was stable during postnatal development, indicating synchronized synaptogenesis. Based on the pure quantity of GABAergic synapses onto interneurons, we propose that mutual inhibition must play an important, yet largely neglected, computational role in the cerebellar cortex.     

Sex‐specific alterations in GABA receptor‐mediated responses in laterodorsal tegmentum are associated with prenatal exposure to nicotine

Smoking during pregnancy is associated with deleterious physiological and cognitive effects on the offspring, which are likely due to nicotine‐induced alteration in the development of neurotransmitter systems. Prenatal nicotine exposure (PNE) in rodents is associated with changes in behaviors controlled in part by the pontine laterodorsal tegmentum (LDT), and LDT excitatory signaling is altered in a sex and age‐dependent manner by PNE. As effects on GABAergic LDT signaling are unknown, we used calcium imaging to evaluate GABA_A receptor‐ (GABA_AR as well as GABA_A‐_ρR) and GABA_B receptor (GABA_BR)‐mediated calcium responses in LDT brain slices from female and male PNE mice in two different age groups. Overall, in older PNE females, changes in calcium induced by stimulation of GABA_AR and GABA_BR, including GABA_A‐_ρR were shifted toward calcium rises. In both young and old males, PNE was associated with alterations in calcium mediated by all three receptors; however, the GABA_AR was the most affected. These results show for the first time that PNE is associated with alterations in GABAergic transmission in the LDT in a sex‐ and age‐dependent manner, and these data are the first to show PNE‐associated alterations in functionality of GABA receptors in any nucleus. PNE‐associated alterations in LDT GABAergic transmission within the LDT would be expected to alter output to target regions and could play a role in LDT‐implicated, negative behavioral outcomes following gestational exposure to smoking. Accordingly, our data provide further supportive evidence of the importance of eliminating the consumption of nicotine during pregnancy.     

Physiological characteristics of NG2-expressing glial cells

Antibodies against the chondroitin sulfate proteoglycan NG2 label a subpopulation of glial cells within the CNS, which have a small cell body and thin radiating processes. Physiological recordings from these small cells in acute brain slices have revealed that they possess unique properties, suggesting that they may comprise a class of glial cells distinct from astrocytes, oligodendrocytes, or microglia. NG2-expressing glial cells (abbreviated as “NG2 cells” here) have a moderate input resistance and are not dye- or tracer-coupled to adjacent cells. They express voltage-gated Na⁺, K⁺and Ca²⁺conductances, though they do not exhibit regenerative Na⁺or Ca²⁺action potentials due to the much larger K⁺conductances present. In addition to voltage-gated conductances, they express receptors for various neurotransmitters. In the hippocampus, AMPA and GABA_Areceptors on these cells are activated by release of transmitter from neurons at defined synaptic junctions that are formed with CA3 pyramidal neurons and GABAergic interneurons. These rapid forms of neuron-glial communication may regulate the proliferation rate of NG2 cells or their development into mature oligodendrocytes. These depolarizing inputs may also trigger the release of neuroactive substances from NG2 cells, providing feedback regulation of signaling at neuronal synapses. Although the presence of Ca²⁺permeable AMPA receptors provides a pathway to link neuronal activity to Ca²⁺dependent processes within the NG2 cells, these receptors also put these cells at risk for glutamate-associated excitotoxicity. This vulnerability to the sustained elevation of glutamate may underlie ischemic induced damage to white matter tracts and contribute to cerebral palsy in premature infants.     

Sustained Glutamate Receptor Activation Down-regulates GABAB Receptors by Shifting the Balance from Recycling to Lysosomal Degradation

Metabotropic GABA_B receptors are abundantly expressed at glutamatergic synapses where they control excitability of the synapse. Here, we tested the hypothesis that glutamatergic neurotransmission may regulate GABA_B receptors. We found that application of glutamate to cultured cortical neurons led to rapid down-regulation of GABA_B receptors via lysosomal degradation. This effect was mimicked by selective activation of AMPA receptors and further accelerated by coactivation of group I metabotropic glutamate receptors. Inhibition of NMDA receptors, blockade of L-type Ca²⁺ channels, and removal of extracellular Ca²⁺ prevented glutamate-induced down-regulation of GABA_B receptors, indicating that Ca²⁺ influx plays a critical role. We further established that glutamate-induced down-regulation depends on the internalization of GABA_B receptors. Glutamate did not affect the rate of GABA_B receptor endocytosis but led to reduced recycling of the receptors back to the plasma membrane. Blockade of lysosomal activity rescued receptor recycling, indicating that glutamate redirects GABA_B receptors from the recycling to the degradation pathway. In conclusion, the data indicate that sustained activation of AMPA receptors down-regulates GABA_B receptors by sorting endocytosed GABA_B receptors preferentially to lysosomes for degradation on the expense of recycling. This mechanism may relieve glutamatergic synapses from GABA_B receptor-mediated inhibition resulting in increased synaptic excitability.     

Dynamic modification of dendritic cable properties and synaptic transmission by voltage-gated potassium channels

Computer simulations of a dendrite possessing voltage-sensitive potassium conductances were used to determine the effects of these conductances on synaptic transmission and on the propagation of synaptic signals within the dendritic tree. Potassium conductances had two principal effects on voltage transients generated by current injections or synaptic conductances. Locally (near the source of the transient), voltage-gated potassium channels produced a potassium shunt current that reduced the amplitude of voltage transients generated by depolarizing currents. This shunt current increased as the amplitude of the depolarizing transient increased and so acted to prevent large synaptic transients from reaching levels that would saturate due to a reduction in driving force. In the presence of rapidly activating potassium currents, excitatory synapses produced larger synaptic currents that were more linearly related to synaptic conductance, but these produced smaller voltage transients. The maximum amplitudes of the voltage transients were limited by the voltage sensitivity of the K⁺ conductance and the rate at which it could activate. Sufficiently rapid synaptic currents could outrun the K⁺ conductance and thus achieve high local peak amplitudes. These effects of K⁺ conductances were unrelated to whether they were located on dendrites or not, being related only to their proximity to the source of synaptic current. The second class of effects of K⁺ conductances depended on their alteration of the electrotonic structure of the postsynaptic cell and so were observed only when they were located on postsynaptic dendrites. Voltage-gated K⁺ conductances produced voltage-dependent electrotonic expansion of depolarized dendrites, which had the effect of isolating synaptic inputs on depolarized dendrites from events on the rest of the neuron. Thus, synapses on the same dendrite interacted destructively to a degree much greater than that expected from the classical driving force nonlinearity. Synapses located proximally to a depolarized dendritic region were less effected than those located distally, and the range of the nonlinear interaction between synapses was dependent on the kinetics of activation and deactivation of the conductance. When present in conjunction with rapidly activating dendritic sodium conductance, the potassium conductance sharpened the requirement for spatial and temporal coincidence to produce synaptic boosting by inward currents, and suppressed out-of-synchrony synaptic inputs.     

Translocation of CaMKII to dendritic microtubules supports the plasticity of local synapses

The processing of excitatory synaptic inputs involves compartmentalized dendritic Ca²⁺ oscillations. The downstream signaling evoked by these local Ca²⁺ transients and their impact on local synaptic development and remodeling are unknown. Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) is an important decoder of Ca²⁺ signals and mediator of synaptic plasticity. In addition to its known accumulation at spines, we observed with live imaging the dynamic recruitment of CaMKII to dendritic subdomains adjacent to activated synapses in cultured hippocampal neurons. This localized and transient enrichment of CaMKII to dendritic sites coincided spatially and temporally with dendritic Ca²⁺ transients. We show that it involved an interaction with microtubular elements, required activation of the kinase, and led to localized dendritic CaMKII autophosphorylation. This process was accompanied by the adjacent remodeling of spines and synaptic AMPA receptor insertion. Replacement of endogenous CaMKII with a mutant that cannot translocate within dendrites lessened this activity-dependent synaptic plasticity. Thus, CaMKII could decode compartmental dendritic Ca²⁺ transients to support remodeling of local synapses.     

Musciomol and flunitrazepam binding sites in a subcellular fraction enriched in rat cerebellar glomeruli

In the internal granular layer of the cerebellar cortex the polysynaptic complexes called glomeruli consist mainly of homogeneous populations of glutamatergic and GABAergic synapses, both located on granule cell dendrites. A subcellular fraction enriched in glomeruli was prepared from rat cerebellum, and the distribution of GABA_A and of benzodiazepine binding sites between membranes derived from this fraction (fraction G) and from a total cerebellar homogenate (fraction T) was studied. The benzodiazepine and GABA binding sites were measured by the binding of agonists [³H]flunitrazepam and [³H]muscimol, respectively. The results indicate that both binding sites are present, but only slightly enriched, in the glomerular synapses. We found a muscimol/flunitrazepam binding site ratio of two, which is consistent with the enrichement of muscimol binding sites in the granular layer shown by both autoradiographic with radioactive glutamatergic ligands and in situ hybridization experiments respectively.     

Regulation of GABA Equilibrium Potential by mGluRs in Rat Hippocampal CA1 Neurons

The equilibrium potential for GABA-A receptor mediated currents (E_GABA) in neonatal central neurons is set at a relatively depolarized level, which is suggested to be caused by a low expression of K⁺/Cl^- co-transporter (KCC2) but a relatively high expression of Na⁺-K⁺-Cl^- cotransporter (NKCC1). Theta-burst stimulation (TBS) in stratum radiatum induces a negative shift in E_GABA in juvenile hippocampal CA1 pyramidal neurons. In the current study, the effects of TBS on E_GABA in neonatal and juvenile hippocampal CA1 neurons and the underlying mechanisms were examined. Metabotropic glutamate receptors (mGluRs) are suggested to modulate KCC2 and NKCC1 levels in cortical neurons. Therefore, the involvement of mGluRs in the regulation of KCC2 or NKCC1 activity, and thus E_GABA, following TBS was also investigated. Whole-cell patch recordings were made from Wistar rat hippocampal CA1 pyramidal neurons, in a slice preparation. In neonates, TBS induces a positive shift in E_GABA, which was prevented by NKCC1 antisense but not NKCC1 sense mRNA. (RS)-a-Methyl-4-carboxyphenylglycine (MCPG), a group I and II mGluR antagonist, blocked TBS-induced shifts in both juvenile and neonatal hippocampal neurons. While blockade of mGluR1 or mGluR5 alone could interfere with TBS-induced shifts in E_GABA in neonates, only a combined blockade could do the same in juveniles. These results indicate that TBS induces a negative shift in E_GABA in juvenile hippocampal neurons but a positive shift in neonatal hippocampal neurons via corresponding changes in KCC2 and NKCC1 expressions, respectively. mGluR activation seems to be necessary for both shifts to occur while the specific receptor subtype involved seems to vary.     

Spatial Relationships between GABAergic and Glutamatergic Synapses on the Dendrites of Distinct Types of Mouse Retinal Ganglion Cells across Development

Neuronal output requires a concerted balance between excitatory and inhibitory (I/E) input. Like other circuits, inhibitory synaptogenesis in the retina precedes excitatory synaptogenesis. How then do neurons attain their mature balance of I/E ratios despite temporal offset in synaptogenesis? To directly compare the development of glutamatergic and GABAergic synapses onto the same cell, we biolistically transfected retinal ganglion cells (RGCs) with PSD95CFP, a marker of glutamatergic postsynaptic sites, in transgenic Thy1­YFPγ2 mice in which GABA_A receptors are fluorescently tagged. We mapped YFPγ2 and PSD95CFP puncta distributions on three RGC types at postnatal day P12, shortly before eye opening, and at P21 when robust light responses in RGCs are present. The mature I_GABA/E ratios varied among ON-Sustained (S) A-type, OFF-S A-type, and bistratified direction selective (DS) RGCs. These ratios were attained at different rates, before eye-opening for ON-S and OFF-S A-type, and after eye-opening for DS RGCs. At both ages examined, the I_GABA/E ratio was uniform across the arbors of the three RGC types. Furthermore, measurements of the distances between neighboring PSD95CFP and YFPγ2 puncta on RGC dendrites indicate that their local relationship is established early in development, and cannot be predicted by random organization. These close spatial associations between glutamatergic and GABAergic postsynaptic sites appear to represent local synaptic arrangements revealed by correlative light and EM reconstructions of a single RGC''s dendrites. Thus, although RGC types have different I_GABA/E ratios and establish these ratios at separate rates, the local relationship between excitatory and inhibitory inputs appear similarly constrained across the RGC types studied.     

Synaptic dendritic activity modulates the single synaptic event

Synaptic transmission is the key system for the information transfer and elaboration among neurons. Nevertheless, a synapse is not a standing alone structure but it is a part of a population of synapses inputting the information from several neurons on a specific area of the dendritic tree of a single neuron. This population consists of excitatory and inhibitory synapses the inputs of which drive the postsynaptic membrane potential in the depolarizing (excitatory synapses) or depolarizing (inhibitory synapses) direction modulating in such a way the postsynaptic membrane potential. The postsynaptic response of a single synapse depends on several biophysical factors the most important of which is the value of the membrane potential at which the response occurs. The concurrence in a specific time window of inputs by several synapses located in a specific area of the dendritic tree can, consequently, modulate the membrane potential such to severely influence the single postsynaptic response. The degree of modulation operated by the synaptic population depends on the number of synapses active, on the relative proportion between excitatory and inbibitory synapses belonging to the population and on their specific mean firing frequencies. In the present paper we show results obtained by the simulation of the activity of a single Glutamatergic excitatory synapse under the influence of two different populations composed of the same proportion of excitatory and inhibitory synapses but having two different sizes (total number of synapses). The most relevant conclusion of the present simulations is that the information transferred by the single synapse is not and independent simple transition between a pre- and a postsynaptic neuron but is the result of the cooperation of all the synapses which concurrently try to transfer the information to the postsynaptic neuron in a given time window. This cooperativeness is mainly operated by a simple mechanism of modulation of the postsynaptic membrane potential which influences the amplitude of the different components forming the postsynaptic excitatory response.