The proper functioning of the adult mammalian brain relies on the orchestrated regulation of neural activity by a diverse population of GABA (gamma-aminobutyric acid)-releasing neurons. Until ...recently, our appreciation of GABA-mediated inhibition focused predominantly on the GABA(A) (GABA type A) receptors located at synaptic contacts, which are activated in a transient or 'phasic' manner by GABA that is released from synaptic vesicles. However, there is growing evidence that low concentrations of ambient GABA can persistently activate certain subtypes of GABA(A) receptor, which are often remote from synapses, to generate a 'tonic' conductance. In this review, we consider the distinct roles of synaptic and extrasynaptic GABA receptor subtypes in the control of neuronal excitability.
CB1 cannabinoid receptors (CB1) are located at axon terminals and effectively control synaptic communication and thereby circuit operation widespread in the CNS. Although it is partially uncovered ...how CB1 activation leads to the reduction of synaptic excitation, the mechanisms of the decrease of GABA release upon activation of these cannabinoid receptors remain elusive. To determine the mechanisms underlying the suppression of synaptic transmission by CB1 at GABAergic synapses, we recorded unitary IPSCs (uIPSCs) at cholecystokinin-expressing interneuron-pyramidal cell connections and imaged presynaptic Ca(2+) transients in mouse hippocampal slices. Our results reveal a power function with an exponent of 2.2 between the amplitude of uIPSCs and intrabouton Ca(2+). Altering CB1 function by either increasing endocannabinoid production or removing its tonic activity allowed us to demonstrate that CB1 controls GABA release by inhibiting Ca(2+) entry into presynaptic axon terminals via N-type (Cav2.2) Ca(2+) channels. These results provide evidence for modulation of intrabouton Ca(2+) influx into GABAergic axon terminals by CB1, leading to the effective suppression of synaptic inhibition.
The exact location of an ion channel on the axo-somato-dendritic surface of a nerve cell crucially affects its functional impact. Recent high-resolution immunolocalization experiments examining the ...distribution of GABA and glutamate receptors, voltage-gated potassium and sodium channels and hyperpolarization-activated mixed cation (HCN) channels clearly demonstrate the lack of simple rules concerning their subcellular distribution. For example, the density of HCN1 subunits in pyramidal cells increases 60-fold from soma to distal dendrites but is uniform over the somato-dendritic surface of olfactory bulb external tufted cells and is highest in the axon of cortical and cerebellar basket cells. Such findings highlight the necessity of determining the precise subcellular location and density of each ion channel in every cell type. Here, I suggest that variations in the subcellular distribution of ion channels are previously unrecognized means of increasing neuronal diversity and, thus, the computational power of the brain.
Glutamate is a major neurotransmitter in the brain that acts both through fast ionotropic receptors and through slower metabotropic receptors coupled to G proteins. Both receptors are present ...throughout the somatodendritic domain of neurons as shown by immunohistochemical and patch clamp recording studies. Immunogold labelling revealed a concentration of metabotropic receptors at the edge, but not within the main body of anatomically defined synapses, raising the possibility that ionotropic and metabotropic receptors are segregated. We applied double immunogold labelling to study glutamatergic parallel and climbing fibre synapses in the cerebellar cortex. The ionotropic AMPA type receptors occupy the membrane opposite the release site in the main body of the synaptic junction, whereas the metabotropic receptors are located at the periphery of the same synapses. Furthermore, immunoreactivity for AMPA receptors is at least twice as high in the parallel fibre synapses as in glutamatergic mossy fibre synapses. We suggest that the spatial segregation of ionotropic and metabotropic glutamate receptors permits the differential activation of these receptors according to the amount of glutamate released presynaptically, whereas the different densities of the ionotropic receptor at distinct synapses could allow the same amount of glutamate to evoke fast responses of different magnitude.
Hippocampal pyramidal cells, receiving domain specific GABAergic inputs, express up to 10 different subunits of the $\gamma $-aminobutyric acid type A (GABA$_{\text{A}}$) receptor, but only 3 ...different subunits are needed to form a functional pentameric channel. We have tested the hypothesis that some subunits are selectively located at subsets of GABAergic synapses. The $\alpha $1 has been found in most GABAergic synapses on all postsynaptic domains of pyramidal cells. In contrast, the $\alpha $2 subunit was located only in a subset of synapses on the somata and dendrites, but in most synapses on axon initial segments innervated by axo-axonic cells. The results demonstrate that molecular specialization in the composition of postsynaptic GABA$_{\text{A}}$ receptor subunits parallels GABAergic cell specialization in targeting synapses to a specific domain of postsynaptic cortical neurons.
Neurones of the olfactory bulb are innervated by GABA-releasing axons and dendrites of diverse origin. Here, I studied GABAergic neurotransmission in juxtaglomerular cells using whole-cell ...voltage-clamp recordings in acute olfactory bulb slices. Spontaneous IPSCs were fully blocked by the GABA
A receptor antagonist SR95531 (40 μM) and the sodium channel blocker tetrodotoxin (1 μM). The IPSCs had mean amplitudes of 125±86 pA and relatively slow biexponential decay times (
τ
1=4.3±1.0 ms (67±12%),
τ
2=16.9±2.7 ms) at physiological temperatures. Short-term plasticity of evoked IPSCs showed two distinct patterns: depressing (
n=4 cells) and facilitating-depressing (
n=9). In two cells, postsynaptic responses were mediated by single functional release sites. During a train of stimuli (4 stimuli at 20 Hz), the release probability increased by two-fold, whereas the potency (postsynaptic responses excluding failures) decreased by ~15%. The increase in release probability for the second stimulus in the train also occurred when the first action potential failed to release transmitter. However, the decrease in the potency was only observed if the preceding action potential released transmitter. These results reveal a heterogeneity in the short-term plasticity of evoked IPSCs in juxtaglomerular cells and demonstrate that the short-term facilitation at some GABAergic synapses is independent of release.
The cellular and subcellular localization of the G1uRA, G1uRB/C and G1uRD subunits of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) type glutamate receptor was determined in the rat ...hippocampus using polyclonal antipeptide antibodies in immunoperoxidase and immunogold procedures. For the localization of the GluRD subunit a new polyclonal antiserum was developed using the C-terminal sequence of the protein (residues 869–881), conjugated to carrier protein and adsorbed to colloidal gold for immunization. The purified antibodies immunoprecipitated about 25% of
3HAMPA binding activity from the hippocampus, cerebellum or whole brain, but very little from neocortex. These antibodies did not precipitate a significant amount of Hkainate binding activity. The antibodies also recognize the G1uRD subunit, but not the other AMPA receptor subunits, when expressed in transfected COS-7 cells and only when permeabilized with detergent, indicating an intracellular epitope.
All subunits were enriched in the neuropil of the dendritic layers of the hippocampus and in the molecular layer of the dentate gyrus. The cellular distribution of the G1uRD subunit was studied more extensively. The strata radiatum, oriens and the dentate molecular layer were more strongly immunoreactive than the stratum lacunosum moleculare, the stratum lucidum and the hilus. However, in the s stratum lucidum of the CA3 area and in the hilus the weakly reacting dendrites were surrounded by immunopositive rosettes, shown in subsequent electron microscopic studies to correspond to complex dendritic spines. In the stratum radiatum, the weakly reacting apical dendrites contrasted with the surrounding intensely stained neuropil. The cell bodies of pyramidal and granule cells were moderately reactive. Some non-principal cells and their dendrites in the pyramidal cell layer and in the alveus also reacted very strongly for the G1uRD subunit.
At the subcellular level, silver intensified immunogold particles for the GluRA, G1uRB/C and GluRD subunits were present at type 1 synaptic membrane specializations on dendritic spines of pyramidal cells throughout all layers of the CAI and CA3 areas. The most densely labelled synapses tended to be on the largest spines and many smaller spines remained unlabelled. Immunoparticle density at type I synapses on dendritic shafts of some non-principal cells was consistently higher than at labelled synapses of dendritic spines of pyramidal cells. Synapses established between dendritic spines and mossy fibre terminals, were immunoreactive for all studied subunits in stratum lucidum of the CA3 area. The postembedding immunogold method revealed that the AMPA type receptors are concentrated within the main body of the anatomically defined type 1 (asymmetrical) synaptic junction. Often only a part of the membrane specialization showed clustered immunoparticles. There was a sharp decrease in immunoreactive receptor density at the edge of the synaptic specialization. Immunolabelling was consistently demonstrated at extrasynaptic sites on dendrites, dendritic spines and somata.
The results demonstrate that the G1uRA, B/C and D subunits of the AMPA type glutamate receptor are present in many of the glutamatergic synapses formed by the entorhinal, CA3 pyramidal and mossy fibre terminals. Some interneurons have a higher density of AMPA type receptors in their asymmetrical afferent synapses than pyramidal cells. This may contribute to a lower activation threshold of interneurons as compared to principal cells by the same afferents in the hippocampal formation.
In some nerve cells, activation of GABA(A) receptors by GABA results in phasic and tonic conductances. Transient activation of synaptic receptors generates phasic inhibition, whereas tonic inhibition ...originates from GABA acting on extrasynaptic receptors, like in cerebellar granule cells, where it is thought to result from the activation of extrasynaptic GABA(A) receptors with a specific subunit composition (alpha(6)beta(x)delta). Here we show that in adult rat hippocampal slices, extracellular GABA levels are sufficiently high to generate a powerful tonic inhibition in delta subunit-expressing dentate gyrus granule cells. In these cells, the mean tonic current is approximately four times larger than that produced by spontaneous synaptic currents occurring at a frequency of approximately 10 Hz. Antagonizing the GABA transporter GAT-1 with NO-711 (2.5 microM) selectively enhanced tonic inhibition by 330% without affecting the phasic component. In contrast, by prolonging the decay of inhibitory postsynaptic currents (IPSCs), the benzodiazepine agonist zolpidem (0.5 microM) augmented phasic inhibition by 66%, while leaving the mean tonic conductance unchanged. These results demonstrate that a tonic GABA(A) receptor-mediated conductance can be recorded from dentate gyrus granule cells of adult rats in in vitro slice preparations. Furthermore, we have identified distinct pharmacological tools to selectively modify tonic and phasic inhibitions, allowing future studies to investigate their specific roles in neuronal function.
In many brain regions, hyperpolarization‐activated cationic currents (Ih) are involved in the generation of rhythmic activities, but the role of Ih in olfactory oscillations remains unclear. ...Knowledge of the cellular and subcellular distributions of hyperpolarization‐activated and cyclic nucleotide‐gated channel (HCN) subunits is necessary for understanding the role of Ih in olfactory network activities. Using light microscopic immunocytochemistry, we demonstrate strong HCN1 labelling of the glomerular layer and moderate staining of granule cell, internal and external plexiform layers of the rat main olfactory bulb. In the glomerular layer, among many unlabelled neurons, two distinct subpopulations of juxtaglomerular cells are labelled. Approximately 10% of the juxtaglomerular cells strongly express HCN1. These small diameter cells are immunoreactive for GABA and comprise a subpopulation of periglomerular cells. An additional subset of juxtaglomerular cells (≈ 1%) expresses low levels of HCN1. They are large in diameter, GABA immunonegative but immunopositive for vesicular glutamate transporter 2, characterizing them as external tufted cells. Quantitative immunogold localization revealed that the somatic plasma membranes of periglomerular cells contain approximately four times more HCN1 labelling than those of external tufted cells. Unlike in cortical pyramidal cells, immunogold density for HCN1 does not significantly differ in somatic and dendritic plasma membranes of external tufted cells, indicating that post‐synaptic potentials arriving at proximal and distal dendrites are modulated by the same density of Ih. Our results demonstrate a cell type‐dependent expression of HCN1 in the olfactory bulb and predict a differential contribution of distinct juxtaglomerular cell types to network oscillations.
Recent technical developments, including antigen-retrieval and electron microscopic immunogold methods, are making it possible to determine some of the basic principles governing the subcellular ...distribution of ionotropic glutamate receptors. Distinct AMPA and NMDA receptor subtypes are selectively targeted to functionally different synapses of a single cell, resulting in an input-selective fine-tuning and regulation of the postsynaptic responses. The amount, density and variability of AMPA receptors at a given glutamatergic synapse is governed by both pre- and postsynaptic factors, resulting in functionally distinct glutamatergic connections that display characteristic patterns of receptor expression.
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